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  • Crystallography, X-Ray  (9)
  • Substrate Specificity  (5)
  • Nature Publishing Group (NPG)  (12)
  • 1
    Publication Date: 2013-09-06
    Description: The ability to design proteins with high affinity and selectivity for any given small molecule is a rigorous test of our understanding of the physiochemical principles that govern molecular recognition. Attempts to rationally design ligand-binding proteins have met with little success, however, and the computational design of protein-small-molecule interfaces remains an unsolved problem. Current approaches for designing ligand-binding proteins for medical and biotechnological uses rely on raising antibodies against a target antigen in immunized animals and/or performing laboratory-directed evolution of proteins with an existing low affinity for the desired ligand, neither of which allows complete control over the interactions involved in binding. Here we describe a general computational method for designing pre-organized and shape complementary small-molecule-binding sites, and use it to generate protein binders to the steroid digoxigenin (DIG). Of seventeen experimentally characterized designs, two bind DIG; the model of the higher affinity binder has the most energetically favourable and pre-organized interface in the design set. A comprehensive binding-fitness landscape of this design, generated by library selections and deep sequencing, was used to optimize its binding affinity to a picomolar level, and X-ray co-crystal structures of two variants show atomic-level agreement with the corresponding computational models. The optimized binder is selective for DIG over the related steroids digitoxigenin, progesterone and beta-oestradiol, and this steroid binding preference can be reprogrammed by manipulation of explicitly designed hydrogen-bonding interactions. The computational design method presented here should enable the development of a new generation of biosensors, therapeutics and diagnostics.〈br /〉〈br /〉〈a href="https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3898436/" target="_blank"〉〈img src="https://static.pubmed.gov/portal/portal3rc.fcgi/4089621/img/3977009" border="0"〉〈/a〉   〈a href="https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3898436/" target="_blank"〉This paper as free author manuscript - peer-reviewed and accepted for publication〈/a〉〈br /〉〈br /〉〈span class="detail_caption"〉Notes: 〈/span〉Tinberg, Christine E -- Khare, Sagar D -- Dou, Jiayi -- Doyle, Lindsey -- Nelson, Jorgen W -- Schena, Alberto -- Jankowski, Wojciech -- Kalodimos, Charalampos G -- Johnsson, Kai -- Stoddard, Barry L -- Baker, David -- P41 GM103533/GM/NIGMS NIH HHS/ -- R01 GM049857/GM/NIGMS NIH HHS/ -- T32 HG000035/HG/NHGRI NIH HHS/ -- T32 HG00035/HG/NHGRI NIH HHS/ -- England -- Nature. 2013 Sep 12;501(7466):212-6. doi: 10.1038/nature12443. Epub 2013 Sep 4.〈br /〉〈span class="detail_caption"〉Author address: 〈/span〉Department of Biochemistry, University of Washington, Seattle, Washington 98195, USA.〈br /〉〈span class="detail_caption"〉Record origin:〈/span〉 〈a href="http://www.ncbi.nlm.nih.gov/pubmed/24005320" target="_blank"〉PubMed〈/a〉
    Keywords: Binding Sites ; Biotechnology ; *Computer Simulation ; Crystallography, X-Ray ; Digoxigenin/chemistry/*metabolism ; *Drug Design ; Estradiol/chemistry/metabolism ; Ligands ; Models, Molecular ; Progesterone/chemistry/metabolism ; Protein Binding ; Proteins/*chemistry/*metabolism ; Reproducibility of Results ; Substrate Specificity
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    Topics: Biology , Chemistry and Pharmacology , Medicine , Natural Sciences in General , Physics
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  • 2
    Publication Date: 2014-10-03
    Description: The CRISPR-associated protein Cas9 is an RNA-guided DNA endonuclease that uses RNA-DNA complementarity to identify target sites for sequence-specific double-stranded DNA (dsDNA) cleavage. In its native context, Cas9 acts on DNA substrates exclusively because both binding and catalysis require recognition of a short DNA sequence, known as the protospacer adjacent motif (PAM), next to and on the strand opposite the twenty-nucleotide target site in dsDNA. Cas9 has proven to be a versatile tool for genome engineering and gene regulation in a large range of prokaryotic and eukaryotic cell types, and in whole organisms, but it has been thought to be incapable of targeting RNA. Here we show that Cas9 binds with high affinity to single-stranded RNA (ssRNA) targets matching the Cas9-associated guide RNA sequence when the PAM is presented in trans as a separate DNA oligonucleotide. Furthermore, PAM-presenting oligonucleotides (PAMmers) stimulate site-specific endonucleolytic cleavage of ssRNA targets, similar to PAM-mediated stimulation of Cas9-catalysed DNA cleavage. Using specially designed PAMmers, Cas9 can be specifically directed to bind or cut RNA targets while avoiding corresponding DNA sequences, and we demonstrate that this strategy enables the isolation of a specific endogenous messenger RNA from cells. These results reveal a fundamental connection between PAM binding and substrate selection by Cas9, and highlight the utility of Cas9 for programmable transcript recognition without the need for tags.〈br /〉〈br /〉〈a href="https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4268322/" target="_blank"〉〈img src="https://static.pubmed.gov/portal/portal3rc.fcgi/4089621/img/3977009" border="0"〉〈/a〉   〈a href="https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4268322/" target="_blank"〉This paper as free author manuscript - peer-reviewed and accepted for publication〈/a〉〈br /〉〈br /〉〈span class="detail_caption"〉Notes: 〈/span〉O'Connell, Mitchell R -- Oakes, Benjamin L -- Sternberg, Samuel H -- East-Seletsky, Alexandra -- Kaplan, Matias -- Doudna, Jennifer A -- P50 GM102706/GM/NIGMS NIH HHS/ -- P50GM102706-03/GM/NIGMS NIH HHS/ -- T32 GM007232/GM/NIGMS NIH HHS/ -- T32 GM066698/GM/NIGMS NIH HHS/ -- Howard Hughes Medical Institute/ -- England -- Nature. 2014 Dec 11;516(7530):263-6. doi: 10.1038/nature13769. Epub 2014 Sep 28.〈br /〉〈span class="detail_caption"〉Author address: 〈/span〉Department of Molecular and Cell Biology, University of California, Berkeley, California 94720, USA. ; Department of Chemistry, University of California, Berkeley, California 94720, USA. ; 1] Howard Hughes Medical Institute, University of California, Berkeley, California 94720, USA [2] Department of Agricultural and Biological Engineering, University of Florida, Gainesville, Florida 32611, USA. ; 1] Department of Molecular and Cell Biology, University of California, Berkeley, California 94720, USA [2] Department of Chemistry, University of California, Berkeley, California 94720, USA [3] Howard Hughes Medical Institute, University of California, Berkeley, California 94720, USA [4] Physical Biosciences Division, Lawrence Berkeley National Laboratory, Berkeley, California 94720, USA.〈br /〉〈span class="detail_caption"〉Record origin:〈/span〉 〈a href="http://www.ncbi.nlm.nih.gov/pubmed/25274302" target="_blank"〉PubMed〈/a〉
    Keywords: Base Sequence ; CRISPR-Associated Proteins/*metabolism ; CRISPR-Cas Systems/*physiology ; Cell Extracts ; Clustered Regularly Interspaced Short Palindromic Repeats/*genetics ; DNA/chemistry/genetics/metabolism ; Genetic Engineering/*methods ; Glyceraldehyde-3-Phosphate Dehydrogenase (Phosphorylating)/genetics ; HeLa Cells ; Humans ; Nucleotide Motifs ; Oligonucleotides/chemistry/genetics/metabolism ; RNA/chemistry/genetics/*metabolism ; RNA, Guide/chemistry/genetics/metabolism ; RNA, Messenger/genetics/isolation & purification/metabolism ; Substrate Specificity
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    Topics: Biology , Chemistry and Pharmacology , Medicine , Natural Sciences in General , Physics
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  • 3
    Publication Date: 2015-12-18
    Description: Tandem repeat proteins, which are formed by repetition of modular units of protein sequence and structure, play important biological roles as macromolecular binding and scaffolding domains, enzymes, and building blocks for the assembly of fibrous materials. The modular nature of repeat proteins enables the rapid construction and diversification of extended binding surfaces by duplication and recombination of simple building blocks. The overall architecture of tandem repeat protein structures--which is dictated by the internal geometry and local packing of the repeat building blocks--is highly diverse, ranging from extended, super-helical folds that bind peptide, DNA, and RNA partners, to closed and compact conformations with internal cavities suitable for small molecule binding and catalysis. Here we report the development and validation of computational methods for de novo design of tandem repeat protein architectures driven purely by geometric criteria defining the inter-repeat geometry, without reference to the sequences and structures of existing repeat protein families. We have applied these methods to design a series of closed alpha-solenoid repeat structures (alpha-toroids) in which the inter-repeat packing geometry is constrained so as to juxtapose the amino (N) and carboxy (C) termini; several of these designed structures have been validated by X-ray crystallography. Unlike previous approaches to tandem repeat protein engineering, our design procedure does not rely on template sequence or structural information taken from natural repeat proteins and hence can produce structures unlike those seen in nature. As an example, we have successfully designed and validated closed alpha-solenoid repeats with a left-handed helical architecture that--to our knowledge--is not yet present in the protein structure database.〈br /〉〈br /〉〈a href="https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4727831/" target="_blank"〉〈img src="https://static.pubmed.gov/portal/portal3rc.fcgi/4089621/img/3977009" border="0"〉〈/a〉   〈a href="https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4727831/" target="_blank"〉This paper as free author manuscript - peer-reviewed and accepted for publication〈/a〉〈br /〉〈br /〉〈span class="detail_caption"〉Notes: 〈/span〉Doyle, Lindsey -- Hallinan, Jazmine -- Bolduc, Jill -- Parmeggiani, Fabio -- Baker, David -- Stoddard, Barry L -- Bradley, Philip -- R01 GM049857/GM/NIGMS NIH HHS/ -- R01 GM115545/GM/NIGMS NIH HHS/ -- R01GM49857/GM/NIGMS NIH HHS/ -- R21 GM106117/GM/NIGMS NIH HHS/ -- R21GM106117/GM/NIGMS NIH HHS/ -- Howard Hughes Medical Institute/ -- England -- Nature. 2015 Dec 24;528(7583):585-8. doi: 10.1038/nature16191. Epub 2015 Dec 16.〈br /〉〈span class="detail_caption"〉Author address: 〈/span〉Division of Basic Sciences, Fred Hutchinson Cancer Research Center, 1100 Fairview Avenue N., Seattle, Washington 98109, USA. ; Department of Biochemistry, University of Washington, Seattle, Washington 98195, USA. ; Institute for Protein Design, University of Washington, Seattle, Washington 98195, USA. ; Howard Hughes Medical Institute, University of Washington, Seattle, Washington 98195, USA. ; Division of Public Health Sciences, Fred Hutchinson Cancer Research Center, 1100 Fairview Avenue N., Seattle, Washington 98019, USA.〈br /〉〈span class="detail_caption"〉Record origin:〈/span〉 〈a href="http://www.ncbi.nlm.nih.gov/pubmed/26675735" target="_blank"〉PubMed〈/a〉
    Keywords: *Amino Acid Motifs ; *Bioengineering ; *Computer Simulation ; Crystallography, X-Ray ; Databases, Protein ; Models, Molecular ; *Protein Structure, Secondary ; Proteins/*chemistry ; Reproducibility of Results
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  • 4
    Publication Date: 2013-05-03
    Description: The smoothened (SMO) receptor, a key signal transducer in the hedgehog signalling pathway, is responsible for the maintenance of normal embryonic development and is implicated in carcinogenesis. It is classified as a class frizzled (class F) G-protein-coupled receptor (GPCR), although the canonical hedgehog signalling pathway involves the GLI transcription factors and the sequence similarity with class A GPCRs is less than 10%. Here we report the crystal structure of the transmembrane domain of the human SMO receptor bound to the small-molecule antagonist LY2940680 at 2.5 A resolution. Although the SMO receptor shares the seven-transmembrane helical fold, most of the conserved motifs for class A GPCRs are absent, and the structure reveals an unusually complex arrangement of long extracellular loops stabilized by four disulphide bonds. The ligand binds at the extracellular end of the seven-transmembrane-helix bundle and forms extensive contacts with the loops.〈br /〉〈br /〉〈a href="https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3657389/" target="_blank"〉〈img src="https://static.pubmed.gov/portal/portal3rc.fcgi/4089621/img/3977009" border="0"〉〈/a〉   〈a href="https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3657389/" target="_blank"〉This paper as free author manuscript - peer-reviewed and accepted for publication〈/a〉〈br /〉〈br /〉〈span class="detail_caption"〉Notes: 〈/span〉Wang, Chong -- Wu, Huixian -- Katritch, Vsevolod -- Han, Gye Won -- Huang, Xi-Ping -- Liu, Wei -- Siu, Fai Yiu -- Roth, Bryan L -- Cherezov, Vadim -- Stevens, Raymond C -- F32 DK088392/DK/NIDDK NIH HHS/ -- P50 GM073197/GM/NIGMS NIH HHS/ -- R01 DA017204/DA/NIDA NIH HHS/ -- R01 DA027170/DA/NIDA NIH HHS/ -- R01 DA27170/DA/NIDA NIH HHS/ -- R01 MH061887/MH/NIMH NIH HHS/ -- R01MH61887/MH/NIMH NIH HHS/ -- U19 MH082441/MH/NIMH NIH HHS/ -- U19 MH82441/MH/NIMH NIH HHS/ -- U54 GM094618/GM/NIGMS NIH HHS/ -- England -- Nature. 2013 May 16;497(7449):338-43. doi: 10.1038/nature12167. Epub 2013 May 1.〈br /〉〈span class="detail_caption"〉Author address: 〈/span〉Department of Integrative Structural and Computational Biology, The Scripps Research Institute, 10550 North Torrey Pines Road, La Jolla, California 92037, USA.〈br /〉〈span class="detail_caption"〉Record origin:〈/span〉 〈a href="http://www.ncbi.nlm.nih.gov/pubmed/23636324" target="_blank"〉PubMed〈/a〉
    Keywords: Amino Acid Motifs ; Amino Acid Sequence ; Antineoplastic Agents/*chemistry/metabolism ; Binding Sites ; Crystallography, X-Ray ; Disulfides/chemistry ; Frizzled Receptors/chemistry/classification ; Humans ; Ligands ; Models, Molecular ; Molecular Sequence Data ; Phthalazines/*chemistry/metabolism ; Protein Structure, Tertiary ; Receptors, G-Protein-Coupled/*chemistry/classification/metabolism ; Structural Homology, Protein
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  • 5
    Publication Date: 2012-03-23
    Description: Opioid receptors mediate the actions of endogenous and exogenous opioids on many physiological processes, including the regulation of pain, respiratory drive, mood, and--in the case of kappa-opioid receptor (kappa-OR)--dysphoria and psychotomimesis. Here we report the crystal structure of the human kappa-OR in complex with the selective antagonist JDTic, arranged in parallel dimers, at 2.9 A resolution. The structure reveals important features of the ligand-binding pocket that contribute to the high affinity and subtype selectivity of JDTic for the human kappa-OR. Modelling of other important kappa-OR-selective ligands, including the morphinan-derived antagonists norbinaltorphimine and 5'-guanidinonaltrindole, and the diterpene agonist salvinorin A analogue RB-64, reveals both common and distinct features for binding these diverse chemotypes. Analysis of site-directed mutagenesis and ligand structure-activity relationships confirms the interactions observed in the crystal structure, thereby providing a molecular explanation for kappa-OR subtype selectivity, and essential insights for the design of compounds with new pharmacological properties targeting the human kappa-OR.〈br /〉〈br /〉〈a href="https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3356457/" target="_blank"〉〈img src="https://static.pubmed.gov/portal/portal3rc.fcgi/4089621/img/3977009" border="0"〉〈/a〉   〈a href="https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3356457/" target="_blank"〉This paper as free author manuscript - peer-reviewed and accepted for publication〈/a〉〈br /〉〈br /〉〈span class="detail_caption"〉Notes: 〈/span〉Wu, Huixian -- Wacker, Daniel -- Mileni, Mauro -- Katritch, Vsevolod -- Han, Gye Won -- Vardy, Eyal -- Liu, Wei -- Thompson, Aaron A -- Huang, Xi-Ping -- Carroll, F Ivy -- Mascarella, S Wayne -- Westkaemper, Richard B -- Mosier, Philip D -- Roth, Bryan L -- Cherezov, Vadim -- Stevens, Raymond C -- P50 GM073197/GM/NIGMS NIH HHS/ -- P50 GM073197-08/GM/NIGMS NIH HHS/ -- R01 DA009045/DA/NIDA NIH HHS/ -- R01 DA009045-17/DA/NIDA NIH HHS/ -- R01 DA017204/DA/NIDA NIH HHS/ -- R01 DA017624/DA/NIDA NIH HHS/ -- R01 DA027170/DA/NIDA NIH HHS/ -- U54 GM094618/GM/NIGMS NIH HHS/ -- U54 GM094618-02/GM/NIGMS NIH HHS/ -- Y1-CO-1020/CO/NCI NIH HHS/ -- Y1-GM-1104/GM/NIGMS NIH HHS/ -- England -- Nature. 2012 Mar 21;485(7398):327-32. doi: 10.1038/nature10939.〈br /〉〈span class="detail_caption"〉Author address: 〈/span〉Department of Molecular Biology, The Scripps Research Institute, 10550 North Torrey Pines Road, La Jolla, California 92037, USA.〈br /〉〈span class="detail_caption"〉Record origin:〈/span〉 〈a href="http://www.ncbi.nlm.nih.gov/pubmed/22437504" target="_blank"〉PubMed〈/a〉
    Keywords: Binding Sites ; Crystallography, X-Ray ; Diterpenes, Clerodane/chemistry/metabolism/pharmacology ; Guanidines/chemistry ; Humans ; Models, Molecular ; Morphinans/chemistry ; Mutagenesis, Site-Directed ; Naltrexone/analogs & derivatives/chemistry/metabolism ; Piperidines/*chemistry/pharmacology ; Protein Conformation ; Receptors, Adrenergic, beta-2/chemistry ; Receptors, CXCR4/chemistry/metabolism ; Receptors, Opioid, kappa/*antagonists & inhibitors/*chemistry/genetics/metabolism ; Structure-Activity Relationship ; Tetrahydroisoquinolines/*chemistry/pharmacology
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    Topics: Biology , Chemistry and Pharmacology , Medicine , Natural Sciences in General , Physics
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  • 6
    Publication Date: 2012-05-19
    Description: Members of the opioid receptor family of G-protein-coupled receptors (GPCRs) are found throughout the peripheral and central nervous system, where they have key roles in nociception and analgesia. Unlike the 'classical' opioid receptors, delta, kappa and mu (delta-OR, kappa-OR and mu-OR), which were delineated by pharmacological criteria in the 1970s and 1980s, the nociceptin/orphanin FQ (N/OFQ) peptide receptor (NOP, also known as ORL-1) was discovered relatively recently by molecular cloning and characterization of an orphan GPCR. Although it shares high sequence similarity with classical opioid GPCR subtypes ( approximately 60%), NOP has a markedly distinct pharmacology, featuring activation by the endogenous peptide N/OFQ, and unique selectivity for exogenous ligands. Here we report the crystal structure of human NOP, solved in complex with the peptide mimetic antagonist compound-24 (C-24) (ref. 4), revealing atomic details of ligand-receptor recognition and selectivity. Compound-24 mimics the first four amino-terminal residues of the NOP-selective peptide antagonist UFP-101, a close derivative of N/OFQ, and provides important clues to the binding of these peptides. The X-ray structure also shows substantial conformational differences in the pocket regions between NOP and the classical opioid receptors kappa (ref. 5) and mu (ref. 6), and these are probably due to a small number of residues that vary between these receptors. The NOP-compound-24 structure explains the divergent selectivity profile of NOP and provides a new structural template for the design of NOP ligands.〈br /〉〈br /〉〈a href="https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3356928/" target="_blank"〉〈img src="https://static.pubmed.gov/portal/portal3rc.fcgi/4089621/img/3977009" border="0"〉〈/a〉   〈a href="https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3356928/" target="_blank"〉This paper as free author manuscript - peer-reviewed and accepted for publication〈/a〉〈br /〉〈br /〉〈span class="detail_caption"〉Notes: 〈/span〉Thompson, Aaron A -- Liu, Wei -- Chun, Eugene -- Katritch, Vsevolod -- Wu, Huixian -- Vardy, Eyal -- Huang, Xi-Ping -- Trapella, Claudio -- Guerrini, Remo -- Calo, Girolamo -- Roth, Bryan L -- Cherezov, Vadim -- Stevens, Raymond C -- P50 GM073197/GM/NIGMS NIH HHS/ -- P50 GM073197-08/GM/NIGMS NIH HHS/ -- R01 DA017204/DA/NIDA NIH HHS/ -- R01 DA017204-08/DA/NIDA NIH HHS/ -- R01 DA027170/DA/NIDA NIH HHS/ -- R01 DA027170-03/DA/NIDA NIH HHS/ -- R01 DA27170/DA/NIDA NIH HHS/ -- U54 GM094618/GM/NIGMS NIH HHS/ -- U54 GM094618-02/GM/NIGMS NIH HHS/ -- Y1-CO-1020/CO/NCI NIH HHS/ -- Y1-GM-1104/GM/NIGMS NIH HHS/ -- England -- Nature. 2012 May 16;485(7398):395-9. doi: 10.1038/nature11085.〈br /〉〈span class="detail_caption"〉Author address: 〈/span〉Department of Molecular Biology, The Scripps Research Institute, La Jolla, California 92037, USA.〈br /〉〈span class="detail_caption"〉Record origin:〈/span〉 〈a href="http://www.ncbi.nlm.nih.gov/pubmed/22596163" target="_blank"〉PubMed〈/a〉
    Keywords: Binding Sites ; Biomimetic Materials/*chemistry/metabolism/pharmacology ; Crystallography, X-Ray ; HEK293 Cells ; Humans ; Ligands ; Models, Molecular ; Narcotic Antagonists ; Opioid Peptides/*chemistry/metabolism/pharmacology ; Piperidines/*chemistry/*metabolism/pharmacology ; Protein Conformation ; Receptors, Opioid/*chemistry/*metabolism ; Receptors, Opioid, kappa/chemistry/metabolism ; Spiro Compounds/*chemistry/*metabolism/pharmacology ; Substrate Specificity
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    Topics: Biology , Chemistry and Pharmacology , Medicine , Natural Sciences in General , Physics
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  • 7
    Publication Date: 2014-01-15
    Description: Opioids represent widely prescribed and abused medications, although their signal transduction mechanisms are not well understood. Here we present the 1.8 A high-resolution crystal structure of the human delta-opioid receptor (delta-OR), revealing the presence and fundamental role of a sodium ion in mediating allosteric control of receptor functional selectivity and constitutive activity. The distinctive delta-OR sodium ion site architecture is centrally located in a polar interaction network in the seven-transmembrane bundle core, with the sodium ion stabilizing a reduced agonist affinity state, and thereby modulating signal transduction. Site-directed mutagenesis and functional studies reveal that changing the allosteric sodium site residue Asn 131 to an alanine or a valine augments constitutive beta-arrestin-mediated signalling. Asp95Ala, Asn310Ala and Asn314Ala mutations transform classical delta-opioid antagonists such as naltrindole into potent beta-arrestin-biased agonists. The data establish the molecular basis for allosteric sodium ion control in opioid signalling, revealing that sodium-coordinating residues act as 'efficacy switches' at a prototypic G-protein-coupled receptor.〈br /〉〈br /〉〈a href="https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3931418/" target="_blank"〉〈img src="https://static.pubmed.gov/portal/portal3rc.fcgi/4089621/img/3977009" border="0"〉〈/a〉   〈a href="https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3931418/" target="_blank"〉This paper as free author manuscript - peer-reviewed and accepted for publication〈/a〉〈br /〉〈br /〉〈span class="detail_caption"〉Notes: 〈/span〉Fenalti, Gustavo -- Giguere, Patrick M -- Katritch, Vsevolod -- Huang, Xi-Ping -- Thompson, Aaron A -- Cherezov, Vadim -- Roth, Bryan L -- Stevens, Raymond C -- P50 GM073197/GM/NIGMS NIH HHS/ -- R01 DA017204/DA/NIDA NIH HHS/ -- U19 MH082441/MH/NIMH NIH HHS/ -- U54 GM094618/GM/NIGMS NIH HHS/ -- Y1-CO-1020/CO/NCI NIH HHS/ -- Y1-GM-1104/GM/NIGMS NIH HHS/ -- England -- Nature. 2014 Feb 13;506(7487):191-6. doi: 10.1038/nature12944. Epub 2014 Jan 12.〈br /〉〈span class="detail_caption"〉Author address: 〈/span〉1] Department of Integrative Structural and Computational Biology, The Scripps Research Institute, 10550 North Torrey Pines Road, La Jolla, California 92037, USA [2]. ; 1] National Institute of Mental Health Psychoactive Drug Screening Program and Department of Pharmacology and Division of Chemical Biology and Medicinal Chemistry, University of North Carolina Chapel Hill Medical School, Chapel Hill, North Carolina 27599, USA [2]. ; Department of Integrative Structural and Computational Biology, The Scripps Research Institute, 10550 North Torrey Pines Road, La Jolla, California 92037, USA. ; National Institute of Mental Health Psychoactive Drug Screening Program and Department of Pharmacology and Division of Chemical Biology and Medicinal Chemistry, University of North Carolina Chapel Hill Medical School, Chapel Hill, North Carolina 27599, USA.〈br /〉〈span class="detail_caption"〉Record origin:〈/span〉 〈a href="http://www.ncbi.nlm.nih.gov/pubmed/24413399" target="_blank"〉PubMed〈/a〉
    Keywords: Allosteric Regulation/drug effects/genetics ; Allosteric Site/drug effects/genetics ; Arrestins/metabolism ; Asparagine/genetics/metabolism ; Crystallography, X-Ray ; Humans ; Ligands ; Models, Molecular ; Mutagenesis, Site-Directed ; Naltrexone/analogs & derivatives/chemistry/metabolism/pharmacology ; Narcotic Antagonists/chemistry/metabolism/pharmacology ; Receptors, Opioid, delta/agonists/antagonists & ; inhibitors/*chemistry/genetics/*metabolism ; *Signal Transduction/drug effects ; Sodium/metabolism/pharmacology ; Structure-Activity Relationship
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    Topics: Biology , Chemistry and Pharmacology , Medicine , Natural Sciences in General , Physics
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  • 8
    Publication Date: 2014-12-04
    Description: Horizontal gene transfer allows organisms to rapidly acquire adaptive traits. Although documented instances of horizontal gene transfer from bacteria to eukaryotes remain rare, bacteria represent a rich source of new functions potentially available for co-option. One benefit that genes of bacterial origin could provide to eukaryotes is the capacity to produce antibacterials, which have evolved in prokaryotes as the result of eons of interbacterial competition. The type VI secretion amidase effector (Tae) proteins are potent bacteriocidal enzymes that degrade the cell wall when delivered into competing bacterial cells by the type VI secretion system. Here we show that tae genes have been transferred to eukaryotes on at least six occasions, and that the resulting domesticated amidase effector (dae) genes have been preserved for hundreds of millions of years through purifying selection. We show that the dae genes acquired eukaryotic secretion signals, are expressed within recipient organisms, and encode active antibacterial toxins that possess substrate specificity matching extant Tae proteins of the same lineage. Finally, we show that a dae gene in the deer tick Ixodes scapularis limits proliferation of Borrelia burgdorferi, the aetiologic agent of Lyme disease. Our work demonstrates that a family of horizontally acquired toxins honed to mediate interbacterial antagonism confers previously undescribed antibacterial capacity to eukaryotes. We speculate that the selective pressure imposed by competition between bacteria has produced a reservoir of genes encoding diverse antimicrobial functions that are tailored for co-option by eukaryotic innate immune systems.〈br /〉〈br /〉〈a href="https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4713192/" target="_blank"〉〈img src="https://static.pubmed.gov/portal/portal3rc.fcgi/4089621/img/3977009" border="0"〉〈/a〉   〈a href="https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4713192/" target="_blank"〉This paper as free author manuscript - peer-reviewed and accepted for publication〈/a〉〈br /〉〈br /〉〈span class="detail_caption"〉Notes: 〈/span〉Chou, Seemay -- Daugherty, Matthew D -- Peterson, S Brook -- Biboy, Jacob -- Yang, Youyun -- Jutras, Brandon L -- Fritz-Laylin, Lillian K -- Ferrin, Michael A -- Harding, Brittany N -- Jacobs-Wagner, Christine -- Yang, X Frank -- Vollmer, Waldemar -- Malik, Harmit S -- Mougous, Joseph D -- AI080609/AI/NIAID NIH HHS/ -- AI083640/AI/NIAID NIH HHS/ -- BB/I020012/1/Biotechnology and Biological Sciences Research Council/United Kingdom -- R01 AI080609/AI/NIAID NIH HHS/ -- R01 AI083640/AI/NIAID NIH HHS/ -- Howard Hughes Medical Institute/ -- England -- Nature. 2015 Feb 5;518(7537):98-101. doi: 10.1038/nature13965. Epub 2014 Nov 24.〈br /〉〈span class="detail_caption"〉Author address: 〈/span〉Department of Microbiology, University of Washington School of Medicine, Seattle, Washington 98195, USA. ; 1] Division of Basic Sciences, Fred Hutchinson Cancer Research Center, Seattle, Washington 98109, USA [2] Howard Hughes Medical Institute, Fred Hutchinson Cancer Research Center, Seattle, Washington 98109, USA. ; Centre for Bacterial Cell Biology, Institute for Cell and Molecular Biosciences, Newcastle University, Newcastle upon Tyne NE2 4AX, UK. ; Department of Microbiology and Immunology, Indiana University School of Medicine, Indianapolis, Indiana 46202, USA. ; 1] Microbial Sciences Institute, Yale University, New Haven, Connecticut 06516, USA [2] Howard Hughes Medical Institute, Yale University, New Haven, Connecticut 06516, USA. ; Department of Cellular and Molecular Pharmacology, University of California, San Francisco, California 94158, USA. ; 1] Microbial Sciences Institute, Yale University, New Haven, Connecticut 06516, USA [2] Howard Hughes Medical Institute, Yale University, New Haven, Connecticut 06516, USA [3] Department of Microbial Pathogenesis, Yale University, New Haven, Connecticut 06516, USA [4] Department of Molecular, Cellular, and Developmental Biology, Yale University, New Haven, Connecticut 06516, USA.〈br /〉〈span class="detail_caption"〉Record origin:〈/span〉 〈a href="http://www.ncbi.nlm.nih.gov/pubmed/25470067" target="_blank"〉PubMed〈/a〉
    Keywords: Amidohydrolases/genetics/metabolism/secretion ; Animals ; Bacteria/cytology/*enzymology/*genetics/immunology ; Bacterial Secretion Systems ; Bacterial Toxins/*genetics/metabolism ; Borrelia burgdorferi/cytology/growth & development/immunology ; Cell Wall/metabolism ; Conserved Sequence/genetics ; Eukaryota/*genetics/*immunology/metabolism ; Gene Transfer, Horizontal/*genetics ; Genes, Bacterial/*genetics ; *Immunity, Innate/genetics ; Ixodes/genetics/immunology/metabolism/microbiology ; Phylogeny ; Substrate Specificity
    Print ISSN: 0028-0836
    Electronic ISSN: 1476-4687
    Topics: Biology , Chemistry and Pharmacology , Medicine , Natural Sciences in General , Physics
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  • 9
    Publication Date: 2011-09-23
    Description: Blood-feeding insects such as mosquitoes are efficient vectors of human infectious diseases because they are strongly attracted by body heat, carbon dioxide and odours produced by their vertebrate hosts. Insect repellents containing DEET (N,N-diethyl-meta-toluamide) are highly effective, but the mechanism by which this chemical wards off biting insects remains controversial despite decades of investigation. DEET seems to act both at close range as a contact chemorepellent, by affecting insect gustatory receptors, and at long range, by affecting the olfactory system. Two opposing mechanisms for the observed behavioural effects of DEET in the gas phase have been proposed: that DEET interferes with the olfactory system to block host odour recognition and that DEET actively repels insects by activating olfactory neurons that elicit avoidance behaviour. Here we show that DEET functions as a modulator of the odour-gated ion channel formed by the insect odorant receptor complex. The functional insect odorant receptor complex consists of a common co-receptor, ORCO (ref. 15) (formerly called OR83B; ref. 16), and one or more variable odorant receptor subunits that confer odour selectivity. DEET acts on this complex to potentiate or inhibit odour-evoked activity or to inhibit odour-evoked suppression of spontaneous activity. This modulation depends on the specific odorant receptor and the concentration and identity of the odour ligand. We identify a single amino-acid polymorphism in the second transmembrane domain of receptor OR59B in a Drosophila melanogaster strain from Brazil that renders OR59B insensitive to inhibition by the odour ligand and modulation by DEET. Our data indicate that natural variation can modify the sensitivity of an odour-specific insect odorant receptor to odour ligands and DEET. Furthermore, they support the hypothesis that DEET acts as a molecular 'confusant' that scrambles the insect odour code, and provide a compelling explanation for the broad-spectrum efficacy of DEET against multiple insect species.〈br /〉〈br /〉〈a href="https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3203342/" target="_blank"〉〈img src="https://static.pubmed.gov/portal/portal3rc.fcgi/4089621/img/3977009" border="0"〉〈/a〉   〈a href="https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3203342/" target="_blank"〉This paper as free author manuscript - peer-reviewed and accepted for publication〈/a〉〈br /〉〈br /〉〈span class="detail_caption"〉Notes: 〈/span〉Pellegrino, Maurizio -- Steinbach, Nicole -- Stensmyr, Marcus C -- Hansson, Bill S -- Vosshall, Leslie B -- R01 DC008600/DC/NIDCD NIH HHS/ -- R01 DC008600-02/DC/NIDCD NIH HHS/ -- R01 DC008600-02S1/DC/NIDCD NIH HHS/ -- R01 DC008600-03/DC/NIDCD NIH HHS/ -- R01 DC008600-03S1/DC/NIDCD NIH HHS/ -- R01 DC008600-03S2/DC/NIDCD NIH HHS/ -- R01 DC008600-04/DC/NIDCD NIH HHS/ -- R01 DC008600-05/DC/NIDCD NIH HHS/ -- Howard Hughes Medical Institute/ -- England -- Nature. 2011 Sep 21;478(7370):511-4. doi: 10.1038/nature10438.〈br /〉〈span class="detail_caption"〉Author address: 〈/span〉Laboratory of Neurogenetics and Behaviour, The Rockefeller University, 1230 York Avenue, Box 63, New York, New York 10065, USA.〈br /〉〈span class="detail_caption"〉Record origin:〈/span〉 〈a href="http://www.ncbi.nlm.nih.gov/pubmed/21937991" target="_blank"〉PubMed〈/a〉
    Keywords: Animals ; Avoidance Learning/drug effects ; Brazil ; DEET/*pharmacology ; Drosophila Proteins ; Drosophila melanogaster/classification/genetics/metabolism ; Insect Repellents/*pharmacology ; Ligands ; *Odors ; Olfactory Receptor Neurons/drug effects ; Polymorphism, Genetic/genetics ; Protein Structure, Tertiary ; Receptors, Odorant/chemistry/*genetics/*metabolism ; Species Specificity ; Substrate Specificity
    Print ISSN: 0028-0836
    Electronic ISSN: 1476-4687
    Topics: Biology , Chemistry and Pharmacology , Medicine , Natural Sciences in General , Physics
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  • 10
    Publication Date: 2011-07-01
    Description: Heavy-metal homeostasis and detoxification is crucial for cell viability. P-type ATPases of the class IB (PIB) are essential in these processes, actively extruding heavy metals from the cytoplasm of cells. Here we present the structure of a PIB-ATPase, a Legionella pneumophila CopA Cu(+)-ATPase, in a copper-free form, as determined by X-ray crystallography at 3.2 A resolution. The structure indicates a three-stage copper transport pathway involving several conserved residues. A PIB-specific transmembrane helix kinks at a double-glycine motif displaying an amphipathic helix that lines a putative copper entry point at the intracellular interface. Comparisons to Ca(2+)-ATPase suggest an ATPase-coupled copper release mechanism from the binding sites in the membrane via an extracellular exit site. The structure also provides a framework to analyse missense mutations in the human ATP7A and ATP7B proteins associated with Menkes' and Wilson's diseases.〈br /〉〈span class="detail_caption"〉Notes: 〈/span〉Gourdon, Pontus -- Liu, Xiang-Yu -- Skjorringe, Tina -- Morth, J Preben -- Moller, Lisbeth Birk -- Pedersen, Bjorn Panyella -- Nissen, Poul -- England -- Nature. 2011 Jun 29;475(7354):59-64. doi: 10.1038/nature10191.〈br /〉〈span class="detail_caption"〉Author address: 〈/span〉Centre for Membrane Pumps in Cells and Disease-PUMPKIN, Danish National Research Foundation, Aarhus University, Department of Molecular Biology, Gustav Wieds Vej 10C, DK-8000 Aarhus C, Denmark.〈br /〉〈span class="detail_caption"〉Record origin:〈/span〉 〈a href="http://www.ncbi.nlm.nih.gov/pubmed/21716286" target="_blank"〉PubMed〈/a〉
    Keywords: Adenosine Triphosphatases/genetics ; Bacterial Proteins/*chemistry/*metabolism ; Binding Sites ; Biological Transport ; Calcium ; Cation Transport Proteins/genetics ; Cell Membrane/metabolism ; Copper/*metabolism ; Crystallography, X-Ray ; Cytoplasm/metabolism ; Hepatolenticular Degeneration/genetics ; Humans ; Legionella pneumophila/*chemistry ; Menkes Kinky Hair Syndrome/genetics ; Models, Molecular ; Mutation, Missense/genetics ; Protein Structure, Tertiary ; Sarcoplasmic Reticulum Calcium-Transporting ATPases/chemistry ; Structure-Activity Relationship
    Print ISSN: 0028-0836
    Electronic ISSN: 1476-4687
    Topics: Biology , Chemistry and Pharmacology , Medicine , Natural Sciences in General , Physics
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