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  • 1
    ISSN: 1432-072X
    Keywords: Phylogeny ; Biochemical evolution ; Aromatic biosynthesis ; Acinetobacter ; Regulatory enzymes
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Key enzymes of aromatic amino acid biosynthesis were examined in the genus Acinetobacter. Members of this genus belong to a suprafamilial assemblage of Gram-negative bacteria (denoted Superfamily B) for which a phylogenetic tree based upon oligonucleotide cataloging of 16S rRNA exists. Since the Acinetobacter lineage diverged at an early evolutionary time from other lineages within Superfamily B, an examination of aromatic biosynthesis in members of this genus has supplied improtant clues for the deduction of major evolutionary events leading to the contemporary aromatic pathways that now exist within Superfamily B. Together with Escherichia coli, Pseudomonas aeruginosa and Xanthomonas campestris, four well-spaced lineages have now been studied in comprehensive detail with respect to comparative enzymological features of aromatic amino acid biosynthesis. A. calcoaceticus and A. lwoffii both possess two chorismate mutase isozymes: one a monofunctional isozyme (chorismate mutase-F), and the other (chorismate mutase-P) a component of a bifunctional P-protein (chorismate mutase-prephenate dehydratase). While both P-protein activities were feedback inhibited by l-phenylalanine, the chorismate mutase-P activity was additionally inhibited by prephenate. Likewise, chorismate mutase-F was product inhibited by prephenate. Two isozymes of 3-deoxy-d-arabino-heptulosonate 7-phosphate synthase were detected. The major isozyme (〉95%) was sensitive to feedback inhibition by l-tyrosine, whereas the minor isozyme was apparently insensitive to allosteric control. Prephenate dehydrogenase and arogenate dehydrogenase activities were both detected, but could not be chromatographically resolved. Available evidence favors the existence of a single dehydrogenase enzyme, exhibiting substrate ambiguity for prephenate andl-arogenate. Dehydrogenase activity with either of the latter substrates was specific for NADP+, NAD+ being ineffective. Consideration of the phylogeny of Superfamily-B organisms suggests that the stem ancestor of the Superfamily possessed a single dehydrogenase enzyme having ambiguity for both substrate and pyridine nucleotide cofactor. Since all other members of Superfamily B have NAD+-specific dehydrogenases, specialization for NADP+ must have occurred following the point of Acinetobacter divergence, leading to the dichotomy seen in present-day Superfamily-B organisms.
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  • 2
    ISSN: 1432-072X
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary The basis for the difference between strains 168 (d-tyrosine-sensitive) and 23 (d-tyrosine-resistant) of Bacillus subtilis at the molecular level is that of transport of d-tyrosine into the cell. Strain 23 does not incorporate significant amounts of d-tyrosine into whole cells. A mutant derivative was isolated from strain 23 which had an altered transport system permitting d-tyrosine uptake, a change which also led to inhibition of growth by d-tyrosine. Strain 168 is extremely sensitive to growth inhibition caused by low concentrations of the d-isomer of tyrosine. A mutant derivative of strain 168 selected for its d-tyrosine resistant phenotype had an altered transport system which no longer recognized the d-isomer of tyrosine. These mutants define at least one element of the tyrosine transport system in B. subtilis and provide a convenient phenotype for the eventual location of the chromosal map position.
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  • 3
    ISSN: 1432-2323
    Source: Springer Online Journal Archives 1860-2000
    Topics: Medicine
    Description / Table of Contents: Résumé Il existe chez 5 à 10 % des femmes qui ont apparemment des résultats normaux de biopsies mammaires, des aspects histologiques et cytologiques d'hyperplasie épithéliale atypique (HEA) du sein qui semblent prédire un certain risque de développer un cancer du sein. Ce risque a été estimé à 4–5 fois celui que présentent les femmes du même âge avec les mêmes autres facteurs de risque, mais qui n'ont pas de telles anomalies histo-cytologiques. Ce risque n'est cependant pas constant et semble diminuer avec le temps, rejoignant le même risques qu'ont d'autres femmes aux mêmes âges 10 à 15 ans après la première détection de ces anomalies. Le risque de voir se développer une prolifération sans atypie cellulaire, même extensive ou complexe, n'est que deux fois celui de la population en générale. L'HEA se voit souvent dans la fratrie des familles a cancer du sein, du moins chez les cousines au premier degré. Ce risque, deux fois celui de l'HEA, se situe à 20% environ 10–15 ans après la biopsie, en particulier chez les femmes de la cinquième (40–49) ou au début de la sixième (50) décennie. Ces considérations ont moins d'importance après l'âge de 60 ans. Un traitement par des oestrogènes à de faibles doses après la ménopause ne semble pas augmenter ce risque. Seuls les cancers intracanalaires in situ non comédocarcinomateux peuvent être considérés comme des lésions précancéreuses et ne nécessitent pas de traitement étendu, que nécessitent, par contre, les lésions du type comédocarcinome, même in situ. Des lésions non comédocarcinomateuses peuvent parfois devenir des cancers invasifs en 6 à 10 ans. Il faut les
    Abstract: Resumen Patrones histológicos y citológicos combinados de hiperplasia epitelial atípica (HA) en la glándula mamaria son indicativos de un riesgo de desarrollar cáncer del orden de 5 a 10% en las mujeres con lesiones por lo demás benignas. El riesgo es 4–5 veces mayor que el de mujeres de poblaciones similares que no poseen tales lesiones. Tales riesgos relativos no son estables, y disminuyen 10–15 años luego de la detección, para aproximarse a los riesgos de mujeres de edades comparables. La enfermedad proliferativa sin atipia, no importa qué tan extensa o compleja sea, predice un ligero riesgo mayor, el cual se acerca al doble del de la población de referencia. Hay una fuerte interacción de la HA con la historia familiar de cáncer mamario en por lo menos los familiaries de primer grado. Tal riesgo dobla al riesgo de HA sola, la dobla y se aproxima al 20% a los 10–15 años después de la biopsia, en particular en mujeres en las edades de los 40 y los primero 50 años, pero tales consideraciones son de menor importancia clínica en mujeres de 60 años. Los estrógenos conjugados en dosis bajas, administrados después de la menopausia, no parecieron incrementar el riesgo por encima del que fue identificado mediante patrones histológicos. Sólo el carcinoma in situ de tipo no comedo puede ser considerado como lesión precursora pero que no da lugar al tratamiento más extenso que se recomienda para el tipo más avanzado de comedo carcinoma ductal in situ. Pequeños carcinomas ductales in situ de tipo no comedo pueden resultar en carcinoma invasivo en un periodo de 6 a 10 años. Estos pueden ser tratados con resección local amplia sìn ìrradiación, pudiendose esperar que probablemente no se presentara recurrencia hasta en 8–10 años de seguimiento. Los carcinomas ductales in situ pueden ser lesiones muy extensas, y esta conducta conservadora debe ser reservada para las lesiones menores.
    Notes: Abstract Specific, combined histologic and cytologic patterns of atypical epithelial hyperplasia (AH) in the breast indicate a medically relevant risk of breast cancer development in 5% to 10% of women with otherwise benign biopsies. This risk is four to five times that of similar women without such lesions, that is, women of the same age and at risk for the same period of time. These relative risks are not stable and fall 10 to 15 years after detection, more closely approximating the risks of women of comparable age. Proliferative disease without atypia, no matter how extensive or complex, predicts only a slight elevation of risk, which approaches double that of the reference population. There is a strong interaction of AH with family history of breast cancer in at least a first degree relative. This risk doubles the risk of AH alone and is approximately 20% at 10 to 15 years after biopsy, particularly for women in their forties and early fifties. These considerations are of less clinical importance in women over age 60. Low replacement doses of conjugated estrogen after the menopause do not further elevate risk beyond that identified by histologic patterns. Noncomedo ductal carcinoma in situ may be considered a true precursor lesions; however, it differs significantly in many ways from the more advanced lesion recognized as the comedo type of ductal carcinoma in situ. Small examples of noncomedo ductal carcinoma in situ can eventuate in invasive carcinoma after 6 to 10 years. They may be treated by wide local excision without radiation, with no recurrence up to 8 to 10 years in all likelihood. Ductal carcinoma in situ lesions can be extensive within the breast, and this conservative posture should be reserved for smaller lesions.
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  • 4
    ISSN: 1432-072X
    Keywords: Phylogeny ; Biochemical evolution ; Aromatic pathway ; L-Phenylalanine ; Pseudomonads
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Group I pseudomonads exhibit diversity for L-phenylalanine biosynthesis that is a basis for separation of two subgroups. Subgroup Ib (fluorescent species such as Pseudomonas aeruginosa, P. fluorescens, or P. putida) possesses an unregulated overflow pathway to Lphenylalanine, together with a second, regulated pathway. Subgroup Ia (non-fluorescent species such as P. stutzeri, P. mendocia, or P. alcaligenes) possess only the regulated pathway to L-phenylalanine. Thus, subgroup Ia species lack an unregulated isozyme of chorismate mutase and arogenate dehydratase, enzymes which are thought to divert chorismate to L-phenylalanine under conditions of high carbon input into aromatic biosynthesis. A priori the overflow pathway could have been either lost in subgroup Ia or gained in subgroup Ib. Since Group V pseudomonads (mainly Xanthomonas) are known to branch off from the Group I lineage at a deeper phylogenetic level than the point of divergence for subgroups Ia and Ib, the presence of the overflow pathway in Group V pseudomonads reveals that the overflow pathway must have been lost in the evolution of subgroup Ia. All Group I species possess a bifunctional protein (P-protein) which catalyzes both chorismate mutase and prephenate dehydratase reactions. In subgroup Ia species this highly conserved protein must be the sole source of prephenate to be used for tyrosine biosynthesis. Thus, the channeling action of the P-protein whereby chorismate is committed towards L-phenylalanine formation can be negated by selective feedback inhibition exerted by L-phenylalanine upon the prephenate dehydratase component of the P-protein. Diversion of prephenate molecules under the latter conditions towards L-tyrosine comprises a channel-shuttle mechanism. Such an essential role of the P-protein chorismate mutase for both tyrosine and phenylalanine formation is supported by the observation that stringent phenylalanine auxotrophs lacked the prephenate dehydratase activity but not chorismate mutase activity of the P-protein.
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  • 5
    ISSN: 1432-072X
    Keywords: Phylogeny ; Pseudomonads ; Aromatic biosynthesis ; DAHP synthase ; Phenylalanine hydroxylase
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract The evolution of aromatic amino acid biosynthesis and its regulation is under study in a large assemblage of prokaryotes (Superfamily A) whose phylogenetic arrangement has been constructed on the criterion of oligonucleotide cataloging. One section of this Superfamily consists of a well defined (rRNA homology) cluster denoted as Group III pseudomonads. Pseudomonas acidovorans ATCC 11299a, a Group III member, was chosen for indepth studies of 3-deoxy-d-arabino-heptulosonate 7-phosphate (DAHP) synthase, the initial regulatory enzyme of aromatic biosynthesis. This strain is of particular interest for evolutionary studies of aromatic metabolism because it possesses phenylalanine hydroxylase, an enzyme whose physiological role and distribution among prokaryotes is largely unknown. Although P. acidovorans ATCC 11299a has been of uncertain identity, we now establish it unambiguously as a species of acidovorans by virtue of its 87% DNA homology with P. acidovorans ATCC 15668 (type strain). This result conformed with enzyme patterning studies which placed ATCC 11299a into pseudomonad Group IIIa, a subgroup containing the acidovorans species. Crude extracts of Group III pseudomonads had previously been shown to share, as a common group characteristic, sensitivity of DAHP synthase to feedback inhibition by either l-tyrosine or l-phenylalanine. Detailed studies with partially purified preparations from strain ATCC 11299a revealed the presence of two distinct regulatory isozymes, DAHP synthase-phe and DAHP synthase-tyr. DAHP synthase-tyr is tightly controlled by l-tyrosine with 50% inhibition of activity being achieved at 4.0 μM effector. DAHP synthase-phe is inhibited 50% by 40 μM l-phenylalanine and exhibits dramatic changes in levels of activity, as well as chromatographic elution patterns, in response to dithiothreitol. This two-isozyme pattern of DAHP synthase has not been described previously, although it may prove to be widespread.
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  • 6
    ISSN: 1432-072X
    Keywords: Phylogeny ; Biochemical evolution ; Aromatic biosynthesis ; Azomonas ; Azotobacter ; Pseudomonas ; Regulatory enzymes
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract The evolutionary history of biochemical pathways can be determined in microbial groupings for which phylogenetic trees have been established. This has been demonstrated best in Superfamily B, an assemblage of rRNA homology groups containing lineages that lead to genera such as Escherichia and other enteric microbes, Pseudomonas (Group I), Xanthomonas, Oceanospirillum, and Acinetobacter. The rRNA homology group that defines Group I pseudomonads also includes Azomonas and Azotobacter, but particular dendrogram points of evolutionary divergence for these genera within Superfamily B have not been established. Phylogenetic relationships at such intergeneric levels can be deduced by analysis of aromaticpathway enzyme arrangement and regulation in selected groupings where dynamic evolutionary changes have occurred. A case in point is illustrated by Axomonas insignis, Azotobacter paspali, and Azotobacter vinelandii — a grouping that appears to be homogeneous with respect to the evolutionary state of the aromatic pathway. The conclusion that this phylogenetic cluster diverges from an ancestor common to pseudomonad subgroup Ia (rather than to subgroup Ib) is based upon the absence of chorismate mutase-F and arogenate dehydratase, enzymes making up a twostep pathway of phenylalanine biosynthesis that is absent in subgroup Ia, but present in subgroup Ib. Of further interest, Azomonas insignis and Azotobacter sp. were found to comprise a distinctive and recently evolved sublineage, differing from subgroup Ia species in their loss of a regulatory isozyme of 3-deoxy-d-arabino-heptulosonate 7-phosphate synthase (ADHP synthase-trp) that is subject to feedback inhibition by l-tryptophan. DAHP synthase-trp is an ancient character state of Superfamily B that has been retained during the evolutionary history of most members of this Superfamily.
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  • 7
    ISSN: 1432-072X
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary L-Phenylalanine is a potent inhibitor of growth in a marine species of blue-green bacteria, Agmenellum quadruplicatum. The growth inhibition is reversed by many amino acids when added to the culture medium simultaneously with L-phenylalanine. The most effective L-phenylalanine antagonists are L-tyrosine, L-alanine, L-leucine, L-methionine, L-tryptophan, and L-isoleucine. However, L-tyrosine is the only effective L-phenylalanine antagonist when growth is inhibited by L-phenylalanine for two or more hours prior to addition of an equimolar concentration of the compound tested as an antagonist. Various explanations that could account for inhibition of growth by L-phenylalanine are discussed. Inhibition of growth by L-phenylalanine is not a feature peculiar to the general physiology of blue-green bacteria. For example, the growth of Anacystis nidulans, a fresh water species, was not inhibited by L-phenylalanine, although a different pattern of metabolite sensitivity was found.
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  • 8
    ISSN: 1617-4623
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary Eighteen auxotrophs of Pseudomonas aeruginosa requiring l-tryptophan for growth were isolated following nitrosoguanidine mutagenesis. Mutant blocks for each step of tryptophan biosynthesis were identified by enzymological assay. A regulatory mutant was characterized which was simultaneously constitutive for the gene products of trpA, trpB and trpD. Another class of regulatory mutant appears to synthesize tryptophan synthetase (i.e., trpE and trpF subunits) constitutively. The results implicate three control entities in the pathway of tryptophan biosynthesis: (i) The gene products of trpA, trpB and trpD are repressible by tryptophan, the range of enzyme specific activity varying at least fifty-fold. (ii) No regulation of the trpC gene product could be demonstrated, indicating that its synthesis is constitutive. (iii) The gene products of rpE and trpF are inducible by indoleglycerol 3-phosphate; the magnitude of induction can exceed 100-fold. These results together with some genetic data indicate a general similarity in gene-enzyme relationships between P. aeruginosa and P. putida. A number of specific differences that distinguish the two species are noted. A mutant blocked in the common pathway of aromatic biosynthesis was used to prove that enzymes of tryptophan biosynthesis other than tryptophan synthetase are not inducible by precursors of the common pathway such as chorismate. It is concluded that the concentration of tryptophan that signals total repression of the gene products of trpA, trpB and trpD is lower than the concentrations necessary for maximal feedback inhibition of anthranilate synthetase and for abolition of the induction of tryptophan synthetase.
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  • 9
    ISSN: 1432-0991
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract The enteric lineage of prokaryotes (traditional enteric bacteria,Aeromonas, andAlteromonas) encompasses closely related genera that share many common character states of aromatic amino acid biosynthesis. For example, they uniformly employ the tightly regulated bifunctional P-protein (chorismate mutase: prephenate dehydratase) to forml-phenylalanine via phenylpyruvate. A second, unregulated pathway to phenylalanine, originally termed the overflow pathway inPseudomonas aeruginosa, consists of a monofunctional chorismate mutase (CM-F) and a cyclohexadienyl dehydratase. The evolution of the overflow pathway has been dynamic in the enteric lineage.Serratia marcescens, Erwinia herbicola, Erwinia amylovora, and several otherErwinia species possess an intact pathway.Salmonella, Klebsiella, andErwinia carotovora possess an incomplete overflow pathway, whileEscherichia, Proteus, Aeromonas, andAlteromonas lack it altogether.
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  • 10
    ISSN: 1432-0991
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract We have isolated a chorismate mutase bradytroph (leaky auxotroph) ofAnabaena sp. PCC 7119 (ATCC 29151) as a spontaneous 6-fluorotryptophan-resistant mutant. The decreased chorismate mutase activity resulted in the production of quantities of the phenylalanine and tyrosine that limited rate of growth. 3-Deoxy-d-arabino-heptulosonate 7-phosphate (DAHP) synthase activity in the mutant was elevated more than twofold over the wild-type activity, suggesting derepression of this enzyme. The physiological deregulation of DAHP synthase and the genetic-based deficiency of chorismate mutase promoted an elevated level of intracellular chorismate, which then overwhelmed the competitive inhibition of anthranilate synthase by tryptophan, resulting in the overproduction of tryptophan and indoleglycerolphosphate. The presence of exogenous serine increased the production of tryptophan at the expense of indoleglycerolphosphate. This indicated that the endogenous potential for increasing the amount of serine available for increased tryptophan production is limited.
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