| One functional switch mediates reversible and irreversible inactivation of a herpesvirus protease. | |
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MedLine Citation:
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PMID: 16533039 Owner: NLM Status: MEDLINE |
Abstract/OtherAbstract:
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Distinct mechanisms have evolved to regulate the function of proteolytic enzymes. Viral proteases in particular have developed novel regulatory mechanisms, presumably due to their comparatively rapid life cycles and responses to constant evolutionary pressure. Herpesviruses are a family of human pathogens that require a viral protease with a concentration-dependent zymogen activation involving folding of two alpha-helices and activation of the catalytic machinery, which results in formation of infectious virions. Kaposi's sarcoma-associated herpesvirus protease (KSHV Pr) is unique among the herpesvirus proteases in possessing an autolysis site in the dimer interface, which removes the carboxyl-terminal 27 amino acids comprising an alpha-helix adjacent to the active site. Truncation results in the irreversible loss of dimerization and concomitant inactivation. We characterized the conformational and functional differences between the active dimer, inactive monomer, and inactive truncated protease to determine the different protease regulatory mechanisms that control the KSHV lytic cycle. Circular dichroism revealed a loss of 31% alpha-helicity upon dimer dissociation. Comparison of the full-length and truncated monomers by NMR showed differences in 21% of the protein structure, mainly located adjacent to the dimer interface, with little perturbation of the overall protein upon truncation. Fluorescence polarization and active site labeling, with a transition state mimetic, characterized the functional effects of these conformational changes. Substrate turnover is abolished in both the full-length and truncated monomers; however, substrate binding remained intact. Disruption of the helix 6 interaction with the active site oxyanion loop is therefore used in two independent regulatory mechanisms of proteolytic activity. |
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Authors:
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Anson M Nomura; Alan B Marnett; Nobuhisa Shimba; Volker Dötsch; Charles S Craik |
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Publication Detail:
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Type: Comparative Study; Journal Article; Research Support, N.I.H., Extramural; Research Support, Non-U.S. Gov't |
Journal Detail:
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Title: Biochemistry Volume: 45 ISSN: 0006-2960 ISO Abbreviation: Biochemistry Publication Date: 2006 Mar |
Date Detail:
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Created Date: 2006-03-14 Completed Date: 2006-05-16 Revised Date: 2007-11-14 |
Medline Journal Info:
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Nlm Unique ID: 0370623 Medline TA: Biochemistry Country: United States |
Other Details:
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Languages: eng Pagination: 3572-9 Citation Subset: IM |
Affiliation:
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Department of Pharmaceutical Chemistry, University of California San Francisco, San Francisco, California 94143, USA. |
Export Citation:
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APA/MLA Format Download EndNote Download BibTex |
| MeSH Terms | |
Descriptor/Qualifier:
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Binding Sites Circular Dichroism Dimerization Enzyme Activation / physiology* Humans Magnetic Resonance Spectroscopy Models, Molecular Protein Structure, Secondary Recombinant Proteins / genetics, metabolism Serine Endopeptidases / physiology* |
| Grant Support | |
ID/Acronym/Agency:
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GM-56531/GM/NIGMS NIH HHS |
| Chemical | |
Reg. No./Substance:
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0/Recombinant Proteins; EC 3.4.21.-/Serine Endopeptidases; EC 3.4.21.-/human herpesvirus 8 protease |
From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine
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