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  • 1990-1994  (97,428)
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  • 1991  (97,428)
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  • 1
    facet.materialart.
    Unknown
    New York, NY : Elsevier
    Keywords: Biochemistry ; Enzymes
    Notes: This is a series title, single volumes see link below.
    ISSN: 1557-7988
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  • 2
    facet.materialart.
    Unknown
    Totowa, NJ : Humana Press
    Keywords: Molecular Biology / methods
    Notes: This is a series title, single volumes see link below.
    ISSN: 1940-6029
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  • 3
    Keywords: Leukemia / therapy ; Prognosis
    Notes: Last volumes with varying subtitle and editor.
    ISSN: 0949-7021
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  • 4
    Keywords: Hazardous Substances / toxicity
    Notes: Ceased with vol. 15(1999).
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  • 5
    facet.materialart.
    Unknown
    Königswinter : Petersberg Verlag
    Keywords: Arbeitsgemeinschaft der Großforschungseinrichtungen (Germany) ; Research institutes / Germany ; Research ; Germany
    Notes: Ceased with ed. 1995.
    ISSN: 0935-2236
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  • 6
    Electronic Resource
    Electronic Resource
    Oxford, UK : Blackwell Publishing Ltd
    Computational intelligence 7 (1991), S. 0 
    ISSN: 1467-8640
    Source: Blackwell Publishing Journal Backfiles 1879-2005
    Topics: Computer Science
    Notes: A large number of wording choices naturally occurring in English sentences cannot be accounted for on semantic or syntactic grounds. They represent arbitrary word usages and are termed collocations. In this paper, we show how collocations can enhance the task of lexical selection in language generation. Previous language generation systems were not able to account for collocations for two reasons: they did not have the lexical information in compiled form and the lexicon formalisms available were not able to handle the variations in collocational knowledge. We describe an implemented generator, Cook, which uses a wide range of collocations to produce sentences in the stock market domain. Cook uses a flexible lexicon containing a range of collocations, from idiomatic phrases to word pairs that were compiled automatically from text corpora using a lexicographic tool, Xtract. We show how Cook is able to merge collocations of various types to produce a wide variety of sentences.
    Type of Medium: Electronic Resource
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  • 7
    Electronic Resource
    Electronic Resource
    Oxford, UK : Blackwell Publishing Ltd
    Computational intelligence 7 (1991), S. 0 
    ISSN: 1467-8640
    Source: Blackwell Publishing Journal Backfiles 1879-2005
    Topics: Computer Science
    Notes: Previous work in natural language génération has exploited discourse focus to guide the selection of propositional content and the génération of referring expressions (e.g., pronominalization, définite noun phrase génération). However, there are many other sources of contextual information which can be used to constrain linguistic realization. The realization of certain classes of temporal and spatial référents, for example, can be guided by more detailed models of time and space. Therefore, this article first identifies a number of contextual coordinates or points of référence (known as indexicals in linguistics). Next, we formalize three of these contextual coordinates – topic, time, and space – in a computational model. In doing so, the article describes the use of a Reichenbachian temporal model which is exploited to guide the realization of verb tense and aspect as well as the realization of temporal référents (e.g., temporal connectives and adverbials such as “meanwhile” and “ten minutes later”). The article then describes a spatial model which is used to guide the linguistic realization of spatial référents (e.g., “here,”“there”) and spatial adverbials (e.g., “two miles away”). We discuss the relation between these temporal and spatial models and illustrate their use in generating text from two different application systems. We find that the combined use of topical, temporal, and spatial contextual coordinates enhances the fluency, connectivity, and conciseness of the resulting text.
    Type of Medium: Electronic Resource
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  • 8
    Electronic Resource
    Electronic Resource
    Oxford, UK : Blackwell Publishing Ltd
    Computational intelligence 7 (1991), S. 0 
    ISSN: 1467-8640
    Source: Blackwell Publishing Journal Backfiles 1879-2005
    Topics: Computer Science
    Type of Medium: Electronic Resource
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  • 9
    Electronic Resource
    Electronic Resource
    Oxford, UK : Blackwell Publishing Ltd
    Computational intelligence 7 (1991), S. 0 
    ISSN: 1467-8640
    Source: Blackwell Publishing Journal Backfiles 1879-2005
    Topics: Computer Science
    Notes: The use of multivalued logics for knowledge representation and nonmonotonic reasoning has often been advocated, in particular within the general framework proposed by Ginsberg in his paper “Multivalued logics: a uniform approach to reasoning in artificial intelligence.” His system is based on a multivalued logic with an arbitrary number of truth values classified with respect to two partial orders, a truth order and a knowledge order. This classification is very interesting and gives an intuitive appeal to the framework. In this paper the work by Ginsberg is critically reviewed, pointing out some flaws and ways to overcome them. Moreover, we present some ideas on how to modify the original schema in order to obtain a more semantically well-founded framework.L'utilisation de la Iogique multivalente pour la représentation des connaissances et le raisonnement non monotone a souvent été préconisée, en particulier à l'intérieur du cadre général proposé par Ginsberg dans son article intitulé? Multivalued logics: a uniform approach to reasoning in artificial intelligence ? Son système est basé sur une logique multivalente comportant un nombre arbitraire de valeurs de vérité classées selon deux ordres partiels: un ordre de vérité et un ordre de connaissances. Cette classification est très intéressante et donne un attrait intuitif au cadre. Dans cet article, l'auteur examine le travail de Ginsberg, y relève des lacunes et propose des moyens de les corriger. De plus, il expose certaines idées en vue de modifier le schéma original et ainsi obtenir un meilleur cadre du point de vue de la sémantique.
    Type of Medium: Electronic Resource
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  • 10
    Electronic Resource
    Electronic Resource
    Oxford, UK : Blackwell Publishing Ltd
    Computational intelligence 7 (1991), S. 0 
    ISSN: 1467-8640
    Source: Blackwell Publishing Journal Backfiles 1879-2005
    Topics: Computer Science
    Notes: When trying to understand a speaker's argument, it is necessary to determine what her claim is and what evidence she provides for it. It is necessary, therefore, to be able to recognize evidence relations in terms of the speaker's beliefs. This paper describes an implementation of an evidence oracle, which tests for evidence between statements and builds a model of the speaker based on the evidence relations found. This implementation is intended to be an advance in the development of practical discourse analysis systems, proposing a basis for verifying certain relationships between utterances. Another contribution of the work is a stratified speaker model which allows for varying levels of acceptance of beliefs attributed to the speaker. Integration of the implemented evidence oracle into a full discourse analyser is presented, together with output illustrating the analysis for several sample arguments. Some extensions of this approach for plan inference are also discussed.Lorsque l'on essaie de comprendre l'argument d'un locuteur, il importe de déterminer la nature de sa prétention et le type d'évidence qui l'accompagne. Par conséquent, il est nécessaire de pouvoir distinguer des relations d'évidence les croyances du locuteur. Cet article décrit la mise en oeuvre d'un oracle qui recherche l'évidence entre des énoncés et construit un modèle du locuteur en fonction des relations d'évidence constatées. Cette mise en oeuvre propose une base pour vérifier certaines relations entre des énoncés; elle se veut une contribution au développement d'un système pratique d'analyse du discours. Une autre contribution de cette recherche est l'élaboration d'un modèle de locuteur stratifyé qui tient compte de niveaux variables d'acceptation des croyances attributeés au locuteur. l'intégration de l'oracle d'évidence sous forme d'analyseur de discours est présentée, ainsi que des illustrations de l' analyse de plusieurs arguments types. Une extension de cette approche à l'inférence de plans est également discutée.
    Type of Medium: Electronic Resource
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  • 11
    Electronic Resource
    Electronic Resource
    Oxford, UK : Blackwell Publishing Ltd
    Computational intelligence 7 (1991), S. 0 
    ISSN: 1467-8640
    Source: Blackwell Publishing Journal Backfiles 1879-2005
    Topics: Computer Science
    Notes: This paper considers computer programs that purport to do inference. It finds the essence of their reasoning ability to reside in a particular relationship which exists between the computations described by the texts of those programs and machine-independent formal representations of the theories in which they are said to reason. The principal novelties in the treatment of this question are the use of explicit “abstraction functions” between states of a computation and sets of sentences of a theory, and the notion of global “inference relations” which characterize the set of all inferences of which a system is capable. The central result of the paper is an “arrow-theoretic” characterization of the concept of a knowledge representation, along with the related concept of a knowledge representation realization. The notions of equivalence and subsumption for knowledge representations are defined, as well as equivalence of knowledge representation realizations. Finally, important issues of expression evaluation and control are addressed. The paper concludes with a slogan: “The AI is in the arrows.”Cet article traite des programmes informatiques qui prétendent faire de ľinférence. Ľauteur constate que ľessentiel de leur capacityé de raisonnement réside dans la relation particulière qui existe entre les calculs décrits par les textes de ces programmes et les représentations formelles non liées à un type de machine des théories dans le cadre desquelles ils sont supposés raisonner. Les principales nouveautés dans le traitement de cette question sont ľutilisation de fonctions ?abstraction explicites éntre les etats ?un calcul et les ensembles de phrases ?une théorie, et la notion de relations ?inférence globales qui caractérisent ľensemble de toutes les inférences dont un système est capable. Le principal résultat de cet article est une caractérisation du concept de la représentation des connaissances, ainsi que du concept connexe de la réalisation de la représentation des connaissances. Les notions ?équivalence et de sous-somption pour la représentation des connaissances sont définies, ainsi que ľéquivalence des réalisations de la représentation des connaissances. Enfin, des questions importantes comme ľanalyse et le contrǒle ?expressions sont traitées.
    Type of Medium: Electronic Resource
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  • 12
    Electronic Resource
    Electronic Resource
    Oxford, UK : Blackwell Publishing Ltd
    Computational intelligence 7 (1991), S. 0 
    ISSN: 1467-8640
    Source: Blackwell Publishing Journal Backfiles 1879-2005
    Topics: Computer Science
    Notes: In this paper, we describe a general framework for the generation of referring expressions. Given an intended referent in the domain, domain-dependent mapping rules build a domain-independent level of recoverable semantic structure, which encodes the semantic content to be recovered by the hearer; a second set of mapping rules then builds an abstract syntactic structure, which is unified with a grammar and lexicon to produce a surface noun phrase. Within this framework, (0 we present a detailed description of the algorithms required to build the recoverable semantic structure, and discuss how the problem of infinite recursion can be handled when using relations in descriptions; and (/;') we show how the two levels of representation facilitate the generation of one-anaphoric noun phrases.
    Type of Medium: Electronic Resource
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  • 13
    Electronic Resource
    Electronic Resource
    Oxford, UK : Blackwell Publishing Ltd
    Computational intelligence 7 (1991), S. 0 
    ISSN: 1467-8640
    Source: Blackwell Publishing Journal Backfiles 1879-2005
    Topics: Computer Science
    Notes: This paper analyzes how a spectrum of architectural and structural ideas fit together to provide the required functionality for explanation generation. Several information processing tasks involved in the choice and organization of the content of an explanation are identified. These are best modeled by distinct mechanisms; hence a particular class of hybrid planning architectures most clearly reflects the nature of the explanation task. The architecture is exemplified by a description of an implemented explanation planner. Various implications of the architecture are discussed, including a classification of structuring relations based on their sources and roles in planning; the elimination of goal-posting preconditions from goal refinement operators; and the level at which nondeterminism is handled.
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  • 14
    Electronic Resource
    Electronic Resource
    Oxford, UK : Blackwell Publishing Ltd
    Computational intelligence 7 (1991), S. 0 
    ISSN: 1467-8640
    Source: Blackwell Publishing Journal Backfiles 1879-2005
    Topics: Computer Science
    Notes: In this paper, we analyze the logical definitions of model-based diagnosis recently presented in the literature, and we propose a unified framework (based on the integration of abductive and consistency-based reasoning) in which most of such definitions can be captured. This allows us to single out the existence of a spectrum of alternatives in the logical definition of diagnosis. A lot of attention in the paper is devoted to analyzing the differences among the definitions in the spectrum. In particular, we show that the definitions can be compared on the basis of their restrictive-ness and we relate such a restrictiveness with the completeness of the model of the system to be diagnosed.Dans cet article, les auteurs analysent les définitions logiques de diagnostics basés sur un modèle dont il a été question récemment dans certains ouvrages. Ils proposent un cadre unifié (basé sur l'intégration du raisonnement abductif et du raisonnement basé sur la consistance) à l'interieur duquel la plupart de ces définitions peuvent ětre regroupées. Cette particularité permet de mettre en lumière l'existence d'un spectre d'alternatives dans la définition logique du diagnostic. Cet article accorde une attention toute particulière à l'analyse des différences entre les définitions du spectre. En outre, les auteurs démontrent que les définitions peuvent ětre compareées en fonction de leur caractère restrictif, qui est ensuite mis en relation avec la complétude du modèle du système faisant l'objet d'un diagnostic.
    Type of Medium: Electronic Resource
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  • 15
    Electronic Resource
    Electronic Resource
    Oxford, UK : Blackwell Publishing Ltd
    Computational intelligence 7 (1991), S. 0 
    ISSN: 1467-8640
    Source: Blackwell Publishing Journal Backfiles 1879-2005
    Topics: Computer Science
    Notes: Self-knowledge is a concept that is present in several philosophies. In this article, we consider the issue of whether or not a learning algorithm can in some sense possess self-knowledge. The question is answered affirmatively. Self-learning inductive inference algorithms are taken to be those that learn programs for their own algorithms, in addition to other functions.La connaissance de soi est un concept qui se retrouve dans plusieurs philosophies. Dans cet article, les auteurs s'interrogent à savoir si un algorithme d' apprentissage peut dans une certaine mesure posséder la connaissance de soi. lis apportent une reponse positive a cette question. Les algorithmes d'inference inductive autodidactes sont ceux qui font l'apprentissage de programmes pour leurs propres algorithmes, en plus d' autres fonctions.
    Type of Medium: Electronic Resource
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  • 16
    Electronic Resource
    Electronic Resource
    Oxford, UK : Blackwell Publishing Ltd
    Computational intelligence 7 (1991), S. 0 
    ISSN: 1467-8640
    Source: Blackwell Publishing Journal Backfiles 1879-2005
    Topics: Computer Science
    Notes: Many AI researchers have come to be dissatisfied with approaches to their discipline based on formal logic. Various alternatives are often suggested, including probability theory. This paper investigates the intimate connection between probability theory and various logics. We show that probability theory, broadly conceived, may be used as a formal semantics for virtually any monotonic logic. Thus, rather than being seen as competing, it is more appropriate to view formal logics as very special cases of probability theory, usually special cases that are computationally more tractable than the more general theory. Thus, probability theory and logic should be seen as complementary. Viewing probability theory in this abstract way may help to shed light on various recalcitrant problems in AI.De nombreux chercheurs dans le domaine de l'intelligence artificielle manifestent une certaine insatisfaction vis-à-vis certaines approches basées sur la logique formelle. Diverses solutions sont souvent proposées, y compris la théorie des probabilityés. Cet article analyse la relation intime entre la théorie des probabilités et diverses logiques. Il est démontré que la théorie des probabilityés, conçue de manière générale, peut ětre utilisée comme une sémantique formelle pour presque toute logique monotonique. Au lieu de percevoir les logiques formelles comme étant en opposition, il est plus approprié de les considérer comme des cas trés spéciaux de la théorie des probabilityés, habituellement plus traitables au niveau calcul que la théorie plus générate. Par conséquent, la théorie des probabilityés et la logique doivent ětre percues comme des éléments complémentaires. Le fait de considérer la théorie des probabilityés d'une manière abstraite peut contribuer à la compréhension de divers problèmes ardus dans le domaine de l'intelligence artificielle.
    Type of Medium: Electronic Resource
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  • 17
    Electronic Resource
    Electronic Resource
    Oxford, UK : Blackwell Publishing Ltd
    Computational intelligence 7 (1991), S. 0 
    ISSN: 1467-8640
    Source: Blackwell Publishing Journal Backfiles 1879-2005
    Topics: Computer Science
    Notes: Naturally occurring discourse often contains expressions such as “however,”“as I have stated before,” and “next.” These expressions, denoted meta-comments, carry important information which helps a hearer speed up the comprehension process. In this paper, we present a mechanism for the generation of meta-comments and their incorporation into computer-generated discourse. This mechanism is based on a simple model of the anticipated effect of the messages to be conveyed and of the meta-comments on the hearer's comprehension. It was implemented in a system called FIGMENT, which generates commentaries on the solution of algebraic equations.Le discours naturel contient souvent des expressions comme 〈〈 cependant 〉〉 et 〈〈 comme je ľai dit auparavant 〉〉. Ces expressions, appelées méta-commentaires, contiennent des informations importantes qui aident à accélérer le processus de compréhension de ľ auditeur. Dans cet article, ľ auteur présente un mécanisme de génération de métacommentaires et ? intégration de ceux-ci au discours créé par ordinateur. Ce mécanisme est basé sur un modèle simple de ľeffet anticipé des messages devant ětre transmis et de ľ';effet des méta-commentaires sur la compréhension de ľ auditeur. II a été mis en oeuvre dans un système appele FIGMENT, qui génère des commentaires sur la solution ?équations algébriques.
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  • 18
    Electronic Resource
    Electronic Resource
    Oxford, UK : Blackwell Publishing Ltd
    Computational intelligence 7 (1991), S. 0 
    ISSN: 1467-8640
    Source: Blackwell Publishing Journal Backfiles 1879-2005
    Topics: Computer Science
    Notes: A question-answering program that provides access to a large amount of data will be most useful if it can tailor its answers to each individual user. In particular, a user's level of knowledge about the domain of discourse is an important factor in this tailoring if the answer provided is to be both informative and understandable to the user. In this research, we address the issue of how the user's domain knowledge, or the level of expertise, might affect an answer. We present TAILOR, a flexible computer system that takes into account this knowledge to provide an answer that is appropriate for users with varying levels of expertise (including novices and experts), without requiring an a priori set of user types.Un programme qui donne accès à un système de connaissances à de nombreux et divers utilisateurs serait plus utile s'il pouvait adapter ses réponses pour chaque utilisateur. En particulier, le savoir de l'utilisateur est un facteur important pour que la réponse soit compréhensible et informative. Dans cet article, nous étudions le rǒle de la connaissance de l'utilisateur pour former un texte qui soit approprié pour la personne qui le lira. Nous présentons TAILOR, un système de génération flexible qui est sensitif au niveau de connaissance de l'utilisateur. TAILOR peut générer des textes appropries pour des utilisateurs aux niveaux de connaissances très variés, sans avoir besoin ?un groupe de stéréotypes définis a priori.
    Type of Medium: Electronic Resource
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  • 19
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    Oxford, UK : Blackwell Publishing Ltd
    Teaching statistics 13 (1991), S. 0 
    ISSN: 1467-9639
    Source: Blackwell Publishing Journal Backfiles 1879-2005
    Topics: Mathematics
    Notes: Using as a starting point pre-existing remarks on the discrepancies arising in calculations of statistical quantities from certain formulae because of errors in the means, or the choice of the appropriate formula, the author proposes a scheme by which teachers and students can overcome these difficulties and achieve error-free results.
    Type of Medium: Electronic Resource
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  • 20
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    Oxford, UK : Blackwell Publishing Ltd
    Teaching statistics 13 (1991), S. 0 
    ISSN: 1467-9639
    Source: Blackwell Publishing Journal Backfiles 1879-2005
    Topics: Mathematics
    Notes: This article describes the use of a data set on 128 countries in the classroom. We can perform various statistical analyses on these data, but the most important message lies in questioning the validity of the data collection.
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  • 21
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    Oxford, UK : Blackwell Publishing Ltd
    Teaching statistics 13 (1991), S. 0 
    ISSN: 1467-9639
    Source: Blackwell Publishing Journal Backfiles 1879-2005
    Topics: Mathematics
    Type of Medium: Electronic Resource
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  • 22
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    Oxford, UK : Blackwell Publishing Ltd
    Teaching statistics 13 (1991), S. 0 
    ISSN: 1467-9639
    Source: Blackwell Publishing Journal Backfiles 1879-2005
    Topics: Mathematics
    Notes: Chong Yuen Chow is a Chinese game played with six dice and sixty-three sticks. This gambling game is a nice little game for students to simulate on the computer. The students may then be able to study its statistical properties.
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  • 23
    Electronic Resource
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    Oxford, UK : Blackwell Publishing Ltd
    Teaching statistics 13 (1991), S. 0 
    ISSN: 1467-9639
    Source: Blackwell Publishing Journal Backfiles 1879-2005
    Topics: Mathematics
    Type of Medium: Electronic Resource
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  • 24
    Electronic Resource
    Electronic Resource
    Oxford, UK : Blackwell Publishing Ltd
    Teaching statistics 13 (1991), S. 0 
    ISSN: 1467-9639
    Source: Blackwell Publishing Journal Backfiles 1879-2005
    Topics: Mathematics
    Type of Medium: Electronic Resource
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  • 25
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    Oxford, UK : Blackwell Publishing Ltd
    Teaching statistics 13 (1991), S. 0 
    ISSN: 1467-9639
    Source: Blackwell Publishing Journal Backfiles 1879-2005
    Topics: Mathematics
    Notes: This article indicates how graphwork can be developed systematically in the secondary school.
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  • 26
    Electronic Resource
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    Oxford, UK : Blackwell Publishing Ltd
    Teaching statistics 13 (1991), S. 0 
    ISSN: 1467-9639
    Source: Blackwell Publishing Journal Backfiles 1879-2005
    Topics: Mathematics
    Type of Medium: Electronic Resource
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  • 27
    Electronic Resource
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    Oxford, UK : Blackwell Publishing Ltd
    Teaching statistics 13 (1991), S. 0 
    ISSN: 1467-9639
    Source: Blackwell Publishing Journal Backfiles 1879-2005
    Topics: Mathematics
    Type of Medium: Electronic Resource
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  • 28
    Electronic Resource
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    Oxford, UK : Blackwell Publishing Ltd
    Teaching statistics 13 (1991), S. 0 
    ISSN: 1467-9639
    Source: Blackwell Publishing Journal Backfiles 1879-2005
    Topics: Mathematics
    Type of Medium: Electronic Resource
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  • 29
    Electronic Resource
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    Oxford, UK : Blackwell Publishing Ltd
    Teaching statistics 13 (1991), S. 0 
    ISSN: 1467-9639
    Source: Blackwell Publishing Journal Backfiles 1879-2005
    Topics: Mathematics
    Type of Medium: Electronic Resource
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  • 30
    Electronic Resource
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    Oxford, UK : Blackwell Publishing Ltd
    Teaching statistics 13 (1991), S. 0 
    ISSN: 1467-9639
    Source: Blackwell Publishing Journal Backfiles 1879-2005
    Topics: Mathematics
    Type of Medium: Electronic Resource
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  • 31
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    Oxford, UK : Blackwell Publishing Ltd
    Teaching statistics 13 (1991), S. 0 
    ISSN: 1467-9639
    Source: Blackwell Publishing Journal Backfiles 1879-2005
    Topics: Mathematics
    Type of Medium: Electronic Resource
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  • 32
    ISSN: 0550-3213
    Source: Elsevier Journal Backfiles on ScienceDirect 1907 - 2002
    Topics: Physics
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  • 33
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    Oxford, UK : Blackwell Publishing Ltd
    Molecular microbiology 5 (1991), S. 0 
    ISSN: 1365-2958
    Source: Blackwell Publishing Journal Backfiles 1879-2005
    Topics: Biology , Medicine
    Notes: Both ATP and an electrochemical potential play roles in translocating proteins across the Inner membrane of Escherichia coli. Recent discoveries have dissected the overall transmembrane movement into separate subreactions with different energy requirements, identified a translocation ATPase, and reconstituted both energy-requiring steps of the reaction from purified components. A more refined understanding of the energetics of this fundamental process is beginning to provide answers about the basic issues of how proteins move across the hydrophobic membrane barrier.
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  • 34
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    Oxford, UK : Blackwell Publishing Ltd
    Molecular microbiology 5 (1991), S. 0 
    ISSN: 1365-2958
    Source: Blackwell Publishing Journal Backfiles 1879-2005
    Topics: Biology , Medicine
    Notes: Regulation of transcription occurs at different levels, one being in the presence of sequences specifically recognized by different forms of RNA polymerase, i.e. the promoters. Three different kinds of promoter are defined according, among other things, to their dependence on the growth rate of the cell: the house-keeper' promoter of many metabolic genes, the stringent promoter found at several rRNA and ribosomal protein genes, and the ‘gearbox’ at genes whose products are required at higher relative amounts at lower growth rates. The identified gearbox promoters of Escherichia coli share specific homologies in the -10, -35 and upstream regions. Although there may be different types of gearbox promoters, the -10 sequence of one of these promoters has been found to be essential for functioning as a gearbox. This suggests the existence of specific sigma factors for its transcription. RpoS (KatF) is a likely candidate for being one of these sigma factors. Computer simulation allows us to predict that such sigma factors should, in turn, be expressed following a gearbox mode, which would then imply the existence of self-regulated loops contributing to the expression of some genes of bacterial division. Some bacterial gene products need to be synthesized al fixed amounts per cell
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  • 35
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    Oxford, UK : Blackwell Publishing Ltd
    Molecular microbiology 5 (1991), S. 0 
    ISSN: 1365-2958
    Source: Blackwell Publishing Journal Backfiles 1879-2005
    Topics: Biology , Medicine
    Notes: Many species within the order Actinomycetales contain one or more soluble cytochrome P450 monooxygenases, often substrate-inducible and responsible for a variety of xenobiotic transformations. The individual cytochromes exhibit a relatively broad substrate specificity, and some strains have the capacity to synthesize large amounts of the protein(s) to compensate for low catalytic turnover with some substrates. All three of the Streptomyces cytochromes sequenced to date are exclusive members of one P450 family, CYP105. In several instances, monooxygenase activity arises from induction of a P450 and associated ferredoxin, or of a P450 only, suggesting that some essential electron donor proteins (reductase and ferredoxin) are not coordinately regulated with the cytochrome. The overall properties of these systems suggest an adaptive strategy whose twofold purpose is to maintain a competitive advantage via the production of secondary metabolites, and, whenever possible, to utilize unusual growth substrates by introducing metabolites from these reactions into the more substrate-specific primary metabolic pathways.
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  • 36
    ISSN: 1365-2958
    Source: Blackwell Publishing Journal Backfiles 1879-2005
    Topics: Biology , Medicine
    Notes: Electrotransformation of Rhodococcus fascians by non-replicating plasmids containing a suitable resistance marker resulted in stable transformants by integration of these constructs at various sites in the genome, thereby generating different mutations. Tagged genes could be isolated in Escherichia coli owing to the presence of a CoIE1 replicon and an ampicillin resistance gene in the inserted sequences.Southern analysis and nucleotide sequencing revealed that recombination can occur at defined locations in the plasmid, while no site preference for target sequences could be detected. Low homology between the recombining sequences indicates illegitimate recombination. The specificity of the plasmid sites could be explained by assuming a linear recombination intermediate, generated by cleavage of the transformed plasmid.
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  • 37
    ISSN: 1365-2958
    Source: Blackwell Publishing Journal Backfiles 1879-2005
    Topics: Biology , Medicine
    Notes: Translational fusions between a mutant phoA (lacking its promoter, ribosomal binding site and signal peptide sequence) and Rhizobium‘symbiotic’ genes were isolated. Since these fusions expressed alkaline phosphatase (AP), the product of phoA, the genes into which phoA was inserted apparently specify proteins located in the bacterial periplasm or cell membrane, the compartment in which AP has activity. These genes were psiA and genes upstream of psiA (psiA is required for normal nodule development and strains with multicopy psiA fall to make exopolysaccharide (EPS) and to nodulate). Fusions between phoA and pss (exo) genes, which are required for EPS production, also resulted in the expression of AP indicating that products of these pss genes were located at the cell surface. Using grus fusions to psiA and pssA, we found that the former was expressed in N2-fixing bean root nodules but the latter was not.
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  • 38
    ISSN: 1365-2958
    Source: Blackwell Publishing Journal Backfiles 1879-2005
    Topics: Biology , Medicine
    Notes: The extracellularly secreted endopeptidase elastase (LasB) is regarded as an important virulence factor of Pseudomonas aeruginosa. It has also been implicated in the processing of LasA which enhances elastolytic activity of LasB. In order to Investigate the role of LasB in virulence and LasA processing, a LasB-negative mutant, PA01E, was constructed by insertional mutagenesis of the LasB structural gene, lasB, in P. aeruginosa PAO. An Internal 636 bp lasB fragment of the plasmid pRB1803 was ligated into a derivative of the mobilization vector pSUP201–1. The resulting plasmid, pBRMOB-LasB, was transformed into Escherichia coli and transferred by filter matings to the LasB-positive P. aeruginosa strain, PA01. Plasmid integration in the lasB site of the chromosome was confirmed by Southern blot analysis. Radioimmunoassay and immunoblotting of PA01E supernatant fluids yielded no detectable LasB (〈1 ng ml-1 LasB). The absence of LasB in PA01E was further proven by the inability of its culture supernatant fluid to cleave transferrin or rabbit immunogiobulin G (IgG) after a 72 h incubation. The residual proteolytic activity of PA01E culture supernatant fluid was attributed to alkaline proteinase (Apr), since it was totally inhibited by specific antibodies against Apr. Residual elastolytic activity in culture supernatant fluid of PAO1E was due to the LasA fragment and to the combined action of the LasA fragment with Apr on elastin. The sizes of purified LasA from PA01 and PA01E were identical (22 kDa). These results show that, besides LasB and the LasA fragment, Apr may also act on elastin in the presence of the LasA fragment and that the proteolytic processing of LasA in P. aeruginosa is independent of LasB.
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  • 39
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    Electronic Resource
    Oxford, UK : Blackwell Publishing Ltd
    Molecular microbiology 5 (1991), S. 0 
    ISSN: 1365-2958
    Source: Blackwell Publishing Journal Backfiles 1879-2005
    Topics: Biology , Medicine
    Notes: An invasive Escherichia coli (EIEC) isolate was found to bind basement-membrane laminin in a saturable and time-dependent manner. Excess of unlabelled laminin inhibited the binding of the radioactively labelled protein. Non-invasive E. coli K-12 exhibited only low-level laminin binding but introduction of the virulence-associated plasmid from the EIEC isolate led to high-level binding. Expression of a receptor for laminin on the bacteria was therefore associated with the presence of the virulence plasmid. Scatchard plot analysis Indicated approximately 1000 receptors per bacterial cell, and a Kd of high-affinity binding of 0.5 pM. A laminin-binding protein which correlated with the presence of the plasmid was isolated and characterized. Its sequence of the eight amino-terminal amino acids was identical to that of the LamB protein of E. coli, although the molecular mass of the two in sodium dodecyl sulphate/polyacrylamide gel (SDS-PAGE) appeared to be slightly different. Both proteins reacted in immunoblot assays with polyclonal antisera raised against either protein, and both proteins bound laminin. Southern-blot hybridization analysis established that both the EIEC strain and the K-12 strains with or without the virulence plasmid contained one lamB gene only, and no laminin-binding protein appeared when the virulence plasmid was introduced into bacteria deleted for the lamB gene. On the basis of these results we suggest that native LamB protein of E. coli or a modified variant of it serves as a major receptor for laminin binding and is present at an increased level in invasive E. coli containing the virulence plasmid.
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  • 40
    ISSN: 1365-2958
    Source: Blackwell Publishing Journal Backfiles 1879-2005
    Topics: Biology , Medicine
    Notes: We describe here the cloning, characterization and analysis of the regulation of the ARO2 gene of Saccharomyces cerevisiae, the first reported study of a eukaryotic gene encoding chorismate synthase (E.C. 4.6.1.4). The gene contains an ORF of 1128 bp, encoding a protein with a calculated molecular mass of 40.8 kDa. ARO2 is regulated under the ‘general control system’ for amino acid biosynthesis by the transcriptional activator GCN4 which binds in vitro at three sites within the AR02 promoter.The AR02 gene product is highly similar to its Escherichia coli counterpart, with a 47% identity distributed over the entire length of the peptide. We therefore suggest that the S. cerevisiae chorismate synthase, in contrast to the Neurospora crassa enzyme, but like other chorismate synthases, is a monofunctional peptide, solely possessing chorismate synthase activity.
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  • 41
    ISSN: 1365-2958
    Source: Blackwell Publishing Journal Backfiles 1879-2005
    Topics: Biology , Medicine
    Notes: We have sequenced a gene in the archaebacterium Sulfolobus solfataricus that codes for a protein that shows sequence similarity to the alpha subunit of the signal recognition particle receptor or docking protein In eukaryotes and the product of the ftsY gene in Escherichia coli. Comparison of the Sulfolobus‘docking protein’ with its eukaryotic and eubacterial counterparts showed that the region of highest sequence similarity corresponds to a GTP-binding site. The presence of this gene in archaebacteria suggests that some of the components involved in protein transport have been conserved in the three kingdoms.
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  • 42
    ISSN: 1365-2958
    Source: Blackwell Publishing Journal Backfiles 1879-2005
    Topics: Biology , Medicine
    Notes: The three-dimensional structure of the regular surface protein (p4 symmetry, lattice constant a=b=10.5 nm) of Comamonas acidovorans has been determined to a resolution of about 1.5 nm by means of electron microscopy and image processing. Three-dimensional reconstructions were performed using native outer membranes and artificial two-dimensional crystals of the surface protein, which was selectively solubilized by deoxycholate and recrystallized on carbon films. The two-fold symmetric morphological complex is composed of two identical monomers which are in tight contact with the outer membrane and presumably anchored to it by a small hydrophobic domain.
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  • 43
    Electronic Resource
    Electronic Resource
    Oxford, UK : Blackwell Publishing Ltd
    Molecular microbiology 5 (1991), S. 0 
    ISSN: 1365-2958
    Source: Blackwell Publishing Journal Backfiles 1879-2005
    Topics: Biology , Medicine
    Notes: In previous work, we showed that Candida albicans has two genes, TEF-1 and TEF-2, which encode identical polypeptides for the highly conserved, essential, protein synthesis factor EF-1α (Breviario et al., 1988). This result prompted questions as to whether C. albicans preferentially uses one of the genes over the other and whether both genes are actually translated into protein. Gene-specific sequence differences in the untranslated portion of each gene made it possible to prepare gene-specific oligonucleotide hybridization probes. Results with the probes showed that the relative steady-state mRNA levels of the two genes were equivalent and that the mRNA for each gene was present in active translation complexes.
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  • 44
    ISSN: 1365-2958
    Source: Blackwell Publishing Journal Backfiles 1879-2005
    Topics: Biology , Medicine
    Notes: Entamoeba histolytica cells secrete electron-dense granules (EDGs) that have collagenase activity. To study the possible involvement of calmodulin (CaM) on EDG secretion, the effect of several CaM antagonists (TFP, R24571, W-7, W-5, dibucaine and DL-propranolol) was tested on this cellular function. Except for W-5 and dibucaine, the rest of these compounds inhibited EDG secretion. Transmission electron microscopy of collagen-activated trophozoites showed numerous EDGs located in or near the surface membrane. In contrast, trophozoites incubated with TFP showed no EDGs. Protein kinase C inhibitors (H-7, ML-9) had no effect on EDG secretion, suggesting that CaM antagonists acted by selectively inhibiting CaM. These results suggest that a CaM-dependent process is involved in EDG secretion.
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  • 45
    ISSN: 1365-2958
    Source: Blackwell Publishing Journal Backfiles 1879-2005
    Topics: Biology , Medicine
    Notes: A strain of Desulfovibrio fructosovorans deleted from the hydN [NiFe]hydrogenase structural gene was constructed. A plasmid carrying a 7 kb DNA fragment on which the hydN gene had been replaced by the npt reporter gene (kanamycin-resistant, KnR) was introduced into D. fructosovorans by electroporation. Southern analysis of one KnR clone demonstrated that the hydN gene had been eliminated by marker exchange. This mutant, which was devoid of the [NiFe]hydrogenase gene, still showed a 10% residual hydrogenase activity. Its ability to grow efficiently with H2 as sole energy source is discussed. This is the first report, in a member of the sulphate-reducing bacteria, of a successful transformation and concomitant homologous recombination leading to a fully controlled genotype.
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  • 46
    ISSN: 1365-2958
    Source: Blackwell Publishing Journal Backfiles 1879-2005
    Topics: Biology , Medicine
    Notes: AmpR, the transcriptional regulator for the Citrobacter freundii ampC β-lactamase gene, was purified. The purified AmpR had DNA-binding activity, the same molecular mass (32 kDa) on sodium dodecyl sulphate polyacrylamide gel electrophoresis as previously described, and N-terminal sequencing of the first 15 amino acids was in agreement with that predicted from the nucleotide sequence. Two mutants were isolated that abolish DNA-binding and β-lactamase induction and which map in the amino- and carboxyl-terminal ends of AmpR, respectively. The mutation in the amino terminus (S35F) was located in a helix-turn-helix region showing high homology to other members of the LysR regulator family. Therefore this mutation may directly abolish the contact between AmpR and its operator sequence. It is suggested that the C-terminal mutation (Y264N) affects subunit interactions in AmpR. One constitutive mutant was isolated which mapped in the centre of the ampR gene. This G102E mutant leads to constitutive β-lactamase expression in the absence of both β-lactam inducer and ampG, a gene essential for induction in wild-type enterobacteria. Another mutant protein, D135Y, showed wild-type properties In an ampG* and an ampG::kan background, but could, unlike wild-type AmpR, activate the ampC gene in an ampG1 mutant background. It is thought that ampG1 is a missense mutant. These two types of ampR mutants suggest that activation of ampC transcription is dependent on the conversion of AmpR into a transcriptional activator and that this activation may normally involve interactions with AmpG.
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  • 47
    ISSN: 1365-2958
    Source: Blackwell Publishing Journal Backfiles 1879-2005
    Topics: Biology , Medicine
    Notes: The fibronectin-binding components (fbcs) of two clinical isolates and a culture collection strain of Streptococcus pyogenes have been analysed. Western immunoblotting of bacterial lysates which had been fractionated on polyacrylamide gels revealed trypsin-sensitive fibronectin-binding species. The genes specifying the fbcs were cloned from all three strains and expressed in Escherichia coli using a λ EMBL3 vector. An fbc gene from the culture collection strain was subcloned and expressed in the E. coli expression vector pJLA601, and subjected to deletion analysis. The fibronectin-binding domain was thereby localized within a 40 kDa truncated peptide encoded by the 1000 bp C-terminal region of the gene. Southern hybridization experiments demonstrated that the analysed gene was present in the parental S. pyogenes chromosome, but not in the DNA of fbc expressing λ clones obtained from the two clinical isolates. Further evidence for the existence of at least two different types of fbcs in group A streptococci was provided by Western blot analysis of recombinant phage tysates which revealed a complex series of fibronectin-binding species ranging from 120 to 200 kDa in size and showing strain-dependent variation in their patterns. As was the case with parental streptococcal strains all of the recombinant fbcs were protease-sensitive, and treatment with trypsin or pronase resulted in a total loss of fibronectin-binding activity. Competitive inhibition experiments indicated that lipoteichoic acid was not a significant fbc in the tested streptococcal strains. The biological separation of the principal fbcs of S. pyogenes from lipoteichoic acid, through the cloning and expression of their determinants in the non-lipoteichoic acid-producing organism E. coli K-12, provides unequivocal proof of the protein nature of these components.
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  • 48
    ISSN: 1365-2958
    Source: Blackwell Publishing Journal Backfiles 1879-2005
    Topics: Biology , Medicine
    Notes: The cryptic ilvlH locus of Salmonella typhimurium has genetic information for two distinct subunits of acetohydroxy acid synthase III. We show that the ilvH-encoded subunit is normally translated and the lack of activity is due to early termination of translation within the promoter-proximal ilvl gene. Analysis of the 5’region of the operon led to identification of the promoter and the amino-terminal part of ilvl. Expression of this gene in a mutant producing acetohydroxy acid synthase is due to a transversion which creates a UUA (leucine) codon in the place of a UGA (stop) codon present in position 12 of the wild-type coding region.
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  • 49
    ISSN: 1365-2958
    Source: Blackwell Publishing Journal Backfiles 1879-2005
    Topics: Biology , Medicine
    Notes: Oligonucleotide-directed mutagenesis of ctxB was used to produce mutants of cholera toxin B subunit (CT-B) altered at residues Cys-9, Gly-33, Lys-34, Arg-35, Cys-86 and Trp-88. Mutants were identified phenotypically by radial passive immune haemolysis assays and genotypically by colony hybridization with specific oligonucleotide probes. Mutant CT-B poly-peptides were characterized for immunoreactivity, binding to ganglioside GM1, ability to associate with the A subunit, ability to form holotoxin, and biological activity. Amino acid substitutions that caused decreased binding of mutant CT-B to ganglioside GM1 and abolished toxicity included negatively charged or large hydrophobic residues for Gly-33 and negatively or positively charged residues for Trp-88. Substitution of lysine or arginine for Gly-33 did not affect immunoreactivity or GM1 -binding activity of CT-B but abolished or reduced toxicity of the mutant holotoxins, respectively. Substitutions of Glu or Asp for Arg-35 interfered with formation of holotoxin, but none of the observed substitutions for Lys-34 or Arg-35 affected binding of CT-B to GM1. The Cys-9, Cys-86 and Trp-88 residues were important for establishing or maintaining the native conformation of CT-B or protecting the CT-B polypeptide from rapid degradation in vivo.
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  • 50
    ISSN: 1365-2958
    Source: Blackwell Publishing Journal Backfiles 1879-2005
    Topics: Biology , Medicine
    Notes: In Escherichia coli, the 5 kb mdoA locus is involved in the osmotically controlled biosynthesis of periplasmic membrane-derived oligosaccharides (MDOs). The structure of this locus was analysed by in vitro cassette insertion, transposon mutagenesis, and gene-fusion analysis.A ‘neo’ cassette, derived from the neomycin phosphotransferase II region of transposon Tn5, was inserted into mdoA, borne by a multicopy plasmid. This plasmid was shown to complement two previously described mdoA mutations, depending on the orientation of the exogenous gene. Thus, the gene altered by these mutations could be expressed under the control of the exogenous promoter. Moreover, the ‘neo’ cassette inactivated another, uncharacterlzed, mdo gene, because when this insertion was transferred into the chromosome MDO synthesis was abolished.The existence of a second gene was confirmed by complementation analysis with a collection of Tn1000 insertions into mdoA. Two groups were defined, and the two genes are organized into an operon (mdoGH). This conclusion was reached because Tn1000 insertions in the first gene displayed a polar effect on the expression of the second gene.An active gene fusion was obtained on a multicopy plasmid between the beginning of mdoH and lacZ. The hybrid β-galactosidase activity followed the same osmotically controlled response as that described for of MDO synthesis. This regulation was unaffected by the presence, or absence, of MDOs In the periplasm. Finally, the amount of mdoA-specific mRNAs, determined by dot blot hybridization, decreased when the osmolarity of the growth medium increased.
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  • 51
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    Oxford, UK : Blackwell Publishing Ltd
    Molecular microbiology 5 (1991), S. 0 
    ISSN: 1365-2958
    Source: Blackwell Publishing Journal Backfiles 1879-2005
    Topics: Biology , Medicine
    Notes: To probe the role of the protective antigen (PA) component of anthrax toxin in toxin entry into animals cells, we examined the membrane channel-forming properties and hydrophobicity of intact and trypsin-cleaved forms of the protein at various pH values. At neutral pH neither form caused release of entrapped K+ from unilamellar lipid vesicles. At pH values below 6.0, however, K+ was rapidly released upon addition of either the nicked PA (PAN) or the 63kDa tryptic fragment of PA (PA63), which has been implicated in the toxin entry process. Under the same conditions intact PA exhibited only weak channel-forming activity, and PA20, the complementary tryptic fragment, showed no such activity. Both PA and PA63 exhibited enhanced hydrophobicity at acidic pH values, but the enhancement was greater and the pH threshold higher with PA63. Our findings indicate that proteolytic removal of PA20 from intact PA enables the residual protein, Pasb63, to adopt a conformation at mildly acidic pH values that permits it to insert readily and form channels in membranes. Thus acidic conditions within endocytic vesicles may trigger membrane insertion of PA63, which in turn promotes translocation of ligated effector moieties, edema factor or lethal factor, across the vesicle membrane into the cytosol.
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  • 52
    ISSN: 1365-2958
    Source: Blackwell Publishing Journal Backfiles 1879-2005
    Topics: Biology , Medicine
    Notes: The gene cluster (rfb region) which determines the biosynthesis of the Shigella flexneri O-antigen of the Y serotype specificity was cloned from a S. flexneri serotype 2a strain. Two plasmids, pPM2212 and pPM2213, which conferred O-antigen biosynthesis were generated from separate cosmid clones by deletion with Cla l. These plasmids expressed O-antigen in Escherichia coli K12 like that of the parental strain, as assessed by reactions to antisera in colony and Western immunoblots, sensitivity to bacteriophage Sf6, and by silver staining of lipopolysaccharides separated by sodium dodecyl sulphate/polyacryl-amide gel electrophoresis. These plasmids also mediated O-antigen expression in an E. coli K12 rfb-delete background, indicating that all the necessary genes have been cloned. A detailed restriction map of the region has been constructed and analysis of various subclones has allowed the limits of the coding region for O-antigen biosynthesis to be defined to a maximum of 11kb. Expression of these plasmids demonstrates a novel phenotype associated with control of lipopolysaccharide chain length. The gene(s) responsible maps adjacent to, but separate from, those associated with the biosynthesis of the O-antigen unit. Analysis of plasmid-encoded proteins in minicells and maxicells has facilitated the construction of a physical map. Finally, plasmid pPM-2212 was used to probe a collection of S. flexneri serotypes by Southern hybridization. With the exception of serotype 6, which appears to be unrelated, a similar pattern was found in all serotypes.
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  • 53
    ISSN: 1365-2958
    Source: Blackwell Publishing Journal Backfiles 1879-2005
    Topics: Biology , Medicine
    Notes: The ammo-terminal pro-sequence consisting of 77 amino acid residues is required to guide the folding of secreted subtilisin E, a serine protease, into active, mature enzyme (Ikemura et al., 1987). Furthermore, denatured subtilisin E can be folded to active enzyme in an intermolecular process with the aid of an exogenously added pro-subtilisin E, the active site of which was mutated (Zhu et al., 1989). In this report, we have synthesized the pro-peptide of 77 residues (corresponding to -1 to -77 in the sequence, where residue +1 is the N-terminal amino acid residue of the mature protein), and have found that it could intermolecularly complement the folding of denatured subtilisin E to active enzyme. Furthermore, we have found that the synthetic pro-peptide exhibits specific strong binding to the active mature enzyme by inhibiting it competitively at its active centre with an upper limit to a Ki of 5.4 × 10−7. In contrast, synthetic pro-peptides corresponding to -44 to -77, -1 to 64 and -1 to -43 inhibited the enzyme with Ki values weaker by two orders of magnitude. The results indicate that the sequence extending from -1 to -77 is essential for specificity of interaction, perhaps generating a conformation that accounts for both roles found hitherto, i.e. specific binding to the active centre, and guiding of the refolding to active enzyme. Thus these results suggest that the pro-peptide functions as an intermolecular chaperone.
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  • 54
    ISSN: 1365-2958
    Source: Blackwell Publishing Journal Backfiles 1879-2005
    Topics: Biology , Medicine
    Notes: Aminoglycoside-phosphotransferases contain several conserved amino acid sequence motifs. Using hydroxylamine we have obtained five independent missense mutations within the aphA-2 gene of transposon Tn5. Four of the mutations dramatically reduced antibiotic resistance. Two were identical and included the replacement of His-188 with Tyr. One other resulted from the replacement of Gly-189 with Asp. These three mutations map within the first of the conserved motifs. The replacement of Asp-261 with Asn maps to the third of these structural motifs. A mutation diminishing but not eliminating aminoglycoside resistance resulted from replacement of the conserved Val-36 with Met. By site-directed mutagenesis three additional mutants were obtained: His-188 was replaced with Leu and Ser, and Arg-211 within the second conserved motif was substituted by Gly. All three showed reduced levels of resistance to kanamycin. Our results show that these conserved motifs are essential for the biological activity of aminoglycoside phosphotransferases.
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  • 55
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    Oxford, UK : Blackwell Publishing Ltd
    Molecular microbiology 5 (1991), S. 0 
    ISSN: 1365-2958
    Source: Blackwell Publishing Journal Backfiles 1879-2005
    Topics: Biology , Medicine
    Notes: The yeast 2 μm plasmid is found in the nucleus of almost all Saccharomyces cerevisiae strains. Its replication is very similar to that of chromosomal DNA. Although the plasmid does not encode essential genes it is stably maintained in the yeast population and exhibits only a small, though detectable, loss rate. This stability is achieved by a plasmid-encoded copy-number control system which ensures constant plasmid levels.For the investigation of 2μm replication, a yeast strain that is absolutely dependent on this plasmid was constructed. This was achieved by disruption of the chromosomal CDC9 gene, coding for DNA ligase and providing this essential gene on a 2μm-derived plasmid. This plasmid is absolutely stable under all growth conditions tested. Using the temperature-sensitive mutant allele cdc9-1 we have developed an artificial control system which allows one to change the copy number of 2μm-derived plasmids solely by changing the incubation temperature.
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  • 56
    ISSN: 1365-2958
    Source: Blackwell Publishing Journal Backfiles 1879-2005
    Topics: Biology , Medicine
    Notes: Since Neisseria gonorrhoeae is an obligate pathogen, there is no animal model for identification of virulence factors for this bacterium. An alternative model for assessment of gonococcal virulence is invasion of the adenocarcinoma endometrial cell line, HeclB. Pre-incubation of gonococci with glutaraldehyde-fixed HeclB cells eliminated the six- to eight-hour lag in entry of bacteria into a fresh HeIIB monolayer seen with unpreincubated gonococci or gonococci pre-incubated in tissue-culture medium alone. Gonococci tightly bound to fixed HecIB cells were more Invasive than cells free in the tissue-culture medium, suggesting that actual contact with HecIB cells was required for the enhancement of invasive ability. Chloramphenicol addition during the preincubation prevented the enhanced invasion. Preincubated gonococci were not more adherent to HecIB cells, suggesting that a stage in invasion after binding of gonococci to HecIB cells was enhanced. The enhanced invasion occurred only when gonococci were preincubated with HecIB ceils and not with HEp-2, HeLa, Chang or CHO cells. This eukaryotic cell specificity for induction of enhanced invasion may indicate a role for invasion in gonococcal infection of the endometrium.
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  • 57
    ISSN: 1365-2958
    Source: Blackwell Publishing Journal Backfiles 1879-2005
    Topics: Biology , Medicine
    Notes: A 2.6kb Clal-Bam HI DNA fragment of megaplasmid 2 of Rhizobium meliloti 2011 was found to carry genes involved in exopolysaccharide synthesis and infection of alfalfa nodules. The analysis of the nucleotide sequence of this DNA fragment revealed the existence of two open reading frames (ORFs) running in opposite directions. Plasmid integration mutagenesis showed that these ORFs are organized as two monocistronic transcription units. One of the ORFs represents a new exo gene designated exoZ, which is involved in, but not essential for, the production of acidic exopolysaccharide. However, exoZ is not necessary for nodule formation with alfalfa. The ExoZ protein was found to show homology (23.3%) to the NodX protein of the R. leguminosarum biovar viciae strain TOM, known to be essential for nodulating the primitive Afghanistan pea. The second identified ORF corresponds to the exoB locus. The deduced amino acid sequence of the ExoB protein is homologous (39.6%) to that of the Escherichia coli GalE protein. In R meliloti, exoB codes for a UDP-glucose 4-epimerase. A deficiency in the activity of this enzyme fully accounts for all the multiple carbohydrate defects that have been observed in exoB mutants.
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  • 58
    Electronic Resource
    Electronic Resource
    Oxford, UK : Blackwell Publishing Ltd
    Molecular microbiology 5 (1991), S. 0 
    ISSN: 1365-2958
    Source: Blackwell Publishing Journal Backfiles 1879-2005
    Topics: Biology , Medicine
    Notes: The population of capsulate Haemophilus influenzae is divided into two phylogenetic divisions. Here we show that in division I strains the capsulation (cap) gene cluster lies between direct repeats of a novel insertion sequence (IS)-like element, IS 1016. cap has apparently been mobilized in the chromosome as a compound transposon by IS 1016, and the repeats have provided a molecular substrate for reversible cap gene amplification, with augmentation of capsule production, through unequal homologous recombination. Such amplification has occurred in serotype b strains, but in these a large direct repeat of cap genes has become fixed in the population. We have found a 1.2kb deletion at one end of this duplicated cap b locus, removing most of one copy of the polysaccharide export gene bexA. We have shown that this makes capsulation dependent on preservation of the direct repeat structure in order to avoid recombination-mediated loss of the other copy of bexA. Type b strains with this cap configuration are disseminated worldwide and currently cause nearly all invasive Haemophilus infections, leading us to speculate that the 1.2 kb deletion occurred in an ancestral type b strain and conferred significant biological advantage.
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  • 59
    ISSN: 1365-2958
    Source: Blackwell Publishing Journal Backfiles 1879-2005
    Topics: Biology , Medicine
    Notes: The transformation of Mycobacterium aurum and Mycobacterium smegmatis with the Gram-negative RSF1010-derived cosmid pJRD215 is described. The plasmid is stably maintained in both species and the antibiotic resistance determinants for kanamycin and streptomycin are expressed. Southern blot analysis shows that rearrangements take place both in M. aurum and in M. smegmatis. The use of pJRD215 in mycobacterial cloning systems is discussed.
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  • 60
    Electronic Resource
    Electronic Resource
    Oxford, UK : Blackwell Publishing Ltd
    Molecular microbiology 5 (1991), S. 0 
    ISSN: 1365-2958
    Source: Blackwell Publishing Journal Backfiles 1879-2005
    Topics: Biology , Medicine
    Notes: The overall transcription of DNA has previously been demonstrated to proceed at extremely high levels of accuracy. We review the evidence that the process of transcription is subject to proof-reading in the Hopfield sense. In addition, we speculate that the proof-reading activity associated with transcription is subject to cyclical phase transitions. That is, during periods of low processivity associated with initiation, RNA synthesis is relatively imprecise. The transition to the elongation phase of RNA synthesis, characterized by a shift to high processivity, is accompanied by enhanced proof-reading. A model for the damping of transcriptional errors, based on a PPi-mediated processive pyrophosphorolysis reaction, is discussed in terms of pausing during transcription.
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  • 61
    ISSN: 1365-2958
    Source: Blackwell Publishing Journal Backfiles 1879-2005
    Topics: Biology , Medicine
    Notes: The micronuclear and macronuclear configurations of a gene encoding the protein synthesis elongation factor EF 1α in the hypotrich ciliate Stylonychia lemnae were compared. The two sequences are generally colinear. The coding sequence of the micronuclear gene is, however, interrupted by a 64 bp insert flanked by a 2 bp direct repeat in a gene region which is moderately conserved among EF 1α genes of different organisms. The insertion site is distinct from known intron positions in eukaryotic EF 1α genes. The insert sequence shows inverted repeats at its ends and thus exhibits typical features of an internal eliminated sequence (IES). Comparison with other such sequences in the related organism Oyxtricha nova shows that the IES falls into a new group of such elements. The macronuclear gene exhibits a strikingly limited codon usage, which cannot be simply explained by the overall base composition of the DNA but probably also relates to the very high copy number of the macronuclear gene and the putative high amount of the gene product.
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  • 62
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    Electronic Resource
    Oxford, UK : Blackwell Publishing Ltd
    Molecular microbiology 5 (1991), S. 0 
    ISSN: 1365-2958
    Source: Blackwell Publishing Journal Backfiles 1879-2005
    Topics: Biology , Medicine
    Notes: Genetic studies have identified a number of genes whose products appear to be required for the transport of the group A colicins and the single-stranded DNA of certain filamentous bacteriophages into Escherichia coli. Mutations in these genes allow normal binding of the colicins to their outer-membrane receptors and of the bacteriophage to the tip of specific conjugative pili, but do not allow translocation of the macromolecules to their target. These mutations have been designed‘tolerant’(tol) mutations and the protein products specified by these genes appear to comprise part of a transport system known as the Tol import system. Some of these genes have been isolated, sequenced and their protein products localized to the membranes or periplasm of E. coli. Information is also available regarding the domains of the colicins or phage proteins which interact with the Tol proteins. A preliminary model of the location and possible interactions of the Tol proteins is presented.
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  • 63
    ISSN: 1365-2958
    Source: Blackwell Publishing Journal Backfiles 1879-2005
    Topics: Biology , Medicine
    Notes: A chromosomal locus, lic3, one of several involved in lipopolysaccharide (LPS) biosynthesis by Haemophilus influenzae, was cloned and its DNA sequence determined. Iic3 comprises four closely apposed open reading frames (ORFs). ORF1 includes tandem repeats of the tetramer CAAT and two start codons out of frame with each other are found upstream of the repeats. ORF1 encodes a protein with no known homologues. 0RF2 encodes the UDP-galactose-4-epimerase (galE) gene. ORF3 encodes a hydrophobic protein with no known homologues. 0RF4 encodes the adenylate kinase (adk) gene. A deletion/insertion mutation lacking the 3′ end of ORF1, all of galE, and the 5′ end of ORF3 was constructed in the parent Hib strain (RM7004). These mutants had a galE phenotype, as evidenced by galactose sensitivity, altered LPS when grown in the absence of exogenous galactose, and reduced virulence in infant rats.
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  • 64
    ISSN: 1365-2958
    Source: Blackwell Publishing Journal Backfiles 1879-2005
    Topics: Biology , Medicine
    Notes: The virulence functions of Yersinia enterocolitica include the pYV-encoded Yop proteins and YadA adhesin as well as the chromosome-encoded enterotoxin, Yst. The yop and yadA genes form a temperature-activated regulon controlled by the transcriptional activator VirF. Gene virF, also localized on pYV, is itself thermoinduced in the absence of other pYV genes. The enterotoxin ysf gene is silent in some collection strains including strain W22703. This paper describes two Tn5-Tc1 chromosomal insertion mutants of W22703 transcribing virF, and hence the yop and yadA genes, at low temperature. These mutants also resumed their production of Yst, with its typical temperature dependence. Both mutations were insertions in the same gene called ymoA for‘Yersinia modulator’The cloned ymoA gene fully complemented the two mutations. Several properties of the mutants suggest that ymoA encodes a histone-like protein. According to the nucleic acid sequence, the product of ymoA is an 8064 Da protein rich in aspartic acid (9%), glutamic acid (9%) and lysine (10.5%), but the predicted amino acid sequence shows no similarity with any described histone-like protein. This work supports recent reports which propose a role for DNA topology and bacterial chromatin structure in thermoregulation of virulence functions.
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  • 65
    ISSN: 1365-2958
    Source: Blackwell Publishing Journal Backfiles 1879-2005
    Topics: Biology , Medicine
    Notes: The dipeptide permease (Dpp) is one of three genetically distinct peptide-transport systems in enteric bacteria. Dpp also plays a role in chemotaxis towards peptides. We have devised three selections for dpp mutations based on resistance to toxic peptides (bacilysin, valine-containing peptides, and biala-phos). All dpp mutations mapped to a single chromosomal locus between 77 and 78 min in Salmonella typhimurium and at 79.2 min in Escherichia coli. Expression of dpp was constitutive in both species but the absolute level of expression varied widely between strains. At least in part this difference in expression levels is determined by c/s-acting sequences. The dpp locus of E. coli was cloned. The first gene in the operon, dppA, encodes a periplasmic dipeptide-binding protein (DBP) required for dipeptide transport and chemotaxis. Downstream of dppA are other genes required for transport but not for chemotaxis. The dipeptide-binding protein was found to share 26.5% sequence identity with the periplasmic oligopeptide-binding protein OppA.
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  • 66
    ISSN: 1365-2958
    Source: Blackwell Publishing Journal Backfiles 1879-2005
    Topics: Biology , Medicine
    Notes: The sequence was determined of 6493 nucleotides encompassing the bet genes of Escherichia coli which encode the osmoregulatory choline—glycine betaine pathway. Four open reading frames were identified: betA encoding choline dehydrogenase, a flavoprotein of 61.9kDa; betB encoding betaine aldehyde dehydrogenase (52.6kDa); betT encoding a proton-motive-force-driven, high-affinity transport system for choline (75.8kDa); and betl, capable of encoding a protein of 21.8 kDa, implicated as a repressor involved in choline regulation of the bet genes. Identification of the genes was supported by subcloning, physical mapping of λplac Mu53 insertions, amino acid sequence similarity, or N-terminal amino acid sequencing. The bet genes are tightly spaced, with bet T located upstream of, and transcribed divergently to, the tandemly linked betlBA genes.
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  • 67
    ISSN: 1365-2958
    Source: Blackwell Publishing Journal Backfiles 1879-2005
    Topics: Biology , Medicine
    Notes: Previous work in our laboratory suggested that DNA topology could be implicated in the regulation of the division gene ftsZ. To settle this question, we have selected and characterized mutants in the gyrB gene able to phenotypically suppress the defects of the ftsZ84 mutation. No strict correlation was found between the degree of plasmid DNA relaxation and the level of suppression of the thermosensitivity of the ftsZ84 strain. Interestingly, the class of mutants that shows maximal suppression is substantially unaffected in DNA topology. In addition, the amount of ffsZ-specific mRNA in this class of mutants is comparable to that present in the ftsZ84 strain. These results hint that the ability of these gyrB mutants to correct the effects of the ftsZ84 mutation is largely unrelated to the function of the GyrB (as a part of DNA gyrase) in the control of DNA superhelicity and suggest hitherto unsuspected interaction between the ftsZ and gyrB gene products.
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  • 68
    Electronic Resource
    Electronic Resource
    Oxford, UK : Blackwell Publishing Ltd
    Molecular microbiology 5 (1991), S. 0 
    ISSN: 1365-2958
    Source: Blackwell Publishing Journal Backfiles 1879-2005
    Topics: Biology , Medicine
    Notes: Several in-frame linker insertions have been made in various positions in the faeG gene encoding the K88ab fimbrial subunit protein from porcine enterotoxigenic Escherichia coli. The effects of the linker insertions have been investigated with regard to the ability of the mutated fimbrial subunits to be exported to the surface of the bacterial cell and assembled into a fimbrial structure. The structure/function relationship of the subunit protein is discussed in the light of the phenotypes of the mutations constructed, secondary structure predictions, and hydrophilicity plots.
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  • 69
    ISSN: 1365-2958
    Source: Blackwell Publishing Journal Backfiles 1879-2005
    Topics: Biology , Medicine
    Notes: In vitro studies of bacteriophage Mu transposition have shown that the phage-encoded transposase and repressor bind the same sequences on the phage genome. We attempted to test that prediction in vivo and found that Mu repressor directly inhibits transposition. We also found that, in the absence of repressor, constitutive expression of Mu transposition functions pA and pB is lethal in Escherichia coli strains lysogenic for a mini-Mu and that this is the result of intensive replication of the mini-Mu. These findings have important consequences where such mini-Mus are used as genetic tools. We also tested whether in Erwinia chrysanthemi the effect of transposition functions on a resident mini-Mu was the same as in E. coli. We observed that expression of pA alone was lethal in E. chrysanthemi and that a large fraction of the survivors underwent precise excision of the mini-Mu.
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  • 70
    ISSN: 1365-2958
    Source: Blackwell Publishing Journal Backfiles 1879-2005
    Topics: Biology , Medicine
    Notes: Penicillin resistance in pneumococci is due to the appearance of high molecular-weight penicillin-binding proteins (PBPs) that have reduced affinity for the antibiotic. We have compared the PBX2x genes (pbpX) of one penicillin-susceptible and five penicillin-resistant clinical isolates of Streptococcus pneumoniae isolated from various parts of the world. All of the resistant isolates contained a low-affinity form of PBP 2x. The 2kb region of the two penicillin-susceptible isolates differed at only eight nucleotide sites (0.4%) and resulted in one single amino acid difference in PBP 2x. In contrast, the sequences of the PBP 2x genes from the resistant isolates differed overall from those of the susceptible isolates at between 7 and 18% of nucleotide sites and resulted in between 27 and 86 amino acid substitutions in PBP 2x. The altered PBP 2x genes consisted of regions that were similar to those of susceptible strains (〈3% diverged), alternating with regions that were very different (18–23% diverged). The presence of highly diverged regions within the PBP 2x genes of the resistant isolates contrasts with the uniformity of the sequences of the amylomaltase genes from the same isolates, and with the uniformity of the PBP 2x genes in the two susceptible isolates. It suggests that the altered PBP 2x genes have arisen by localized interspecies recombinational events involving the PBP 2x genes of closely related streptococci, as has been suggested to occur for altered PBP 2b genes (Dowson etal., 1989b). The PBP 2x genes from the resistant isolates could transform the susceptible strain R6 to increased levels of resistance to β-lactam antibiotics. Indicating that the altered forms of PBP 2x in the resistant isolates contribute to their resistance to penicillin.
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  • 71
    ISSN: 1365-2958
    Source: Blackwell Publishing Journal Backfiles 1879-2005
    Topics: Biology , Medicine
    Notes: A prominent 19kDa surface antigen of Legionella pneumophila, cloned in Escherichia coli, was found to be intimately associated with peptidoglycan. The DNA region encoding this antigen was mapped on an 11.9kb plasmid by means of deletion analysis and transposon mutagenesis. PhoA+ gene fusions, generated by TnphoA insertions into this region, confirmed the presence of a gene encoding a secreted protein. PhoA+ transposon insertions were also associated with loss of the 19 kDa antigen in immunoassay s using a monoclonal antibody (mAb1E9) and the replacement of the 19kDa antigen with larger fusion proteins in immunoblots using Legionella immune serum. A 1540bp PstI fragment carrying the gene was sequenced, and the open reading frame encoding the antigen was identified. The gene encodes a polypeptide 176 amino acid residues long and 18913Da in size. The presence of a signal sequence of 22 amino acids with a consensus sequence for cleavage by signal peptidase II indicates that the antigen is a lipoprotein, and striking similarity with peptidoglycan-associated lipoproteins (PALs) from E. coli (51% amino acid homology) and Haemophilus influenzae (55% homology) is noted. We conclude that the 19kDa antigen of L. pneumophila is the structural equivalent of the PAL found in other Gram-negative species and suggest that its post-translational acylation may explain its potency as an immunogen.
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  • 72
    ISSN: 1365-2958
    Source: Blackwell Publishing Journal Backfiles 1879-2005
    Topics: Biology