The effect of mutant peptide cofactors on adenovirus protease activity and virus infection.
Adenoviruses encode a cysteine protease, adenain, required for uncoating and virion maturation. Adenain activity is regulated by an 11-amino-acid peptide cofactor thiol-bonded distal to the active site. Structural and experimental data suggest that the peptide might stabilize adenain in an optimal conformation for enzyme activity by bridging two noncontiguous regions of the molecule. The sequence requirements for this mechanism were examined both in vitro and ex vivo by means of mutant peptides and databank analysis. The results of in vitro experiments suggested that activation is not an all or nothing mechanism. With the exception of the smallest peptide, the mutant peptides bound to adenain, activated it, and competed with the wild-type peptide, but all of this occurred with reduced efficiency. When added to the medium of infected cells, most of the peptides inhibited infectious virus production to varying degrees in a dose-dependent manner and in accordance with their in vitro activity on adenain. We interpret this inhibition to be due to unscheduled adenain activation. Examination of the activation peptide sequences from 19 adenovirus serotypes revealed a limited number of conserved sequence features. These features were in agreement with the experimental data. We conclude that binding and activation of adenain by pVIc may be reversible and this reversibility may be an integral aspect of the in vivo regulation of enzyme activity in the course of virus assembly. The peptide cofactor binding domain is therefore a potential target for the development of anti-adenoviral agents.
Ruzindana-Umunyana
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A
Departement de Microbiologie et d'Infectiologie, Universite de Sherbrooke, Sherbrooke, Quebec, J1H 5N4, Canada.
Sircar
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S
Weber
J M
JM
D000260:Adenoviruses, Human / Q000187:drug effects / Q000201:enzymology* / Q000235:genetics / Q000254:growth & development*; D000595:Amino Acid Sequence; D001665:Binding Sites; D002460:Cell Line; D003067:Coenzymes / Q000737:chemistry / Q000235:genetics / Q000378:metabolism / Q000494:pharmacology*; D017124:Conserved Sequence / Q000235:genetics; D003546:Cysteine Endopeptidases / Q000235:genetics / Q000378:metabolism*; D004220:Disulfides / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D004789:Enzyme Activation / Q000187:drug effects; D004795:Enzyme Stability / Q000187:drug effects; D005453:Fluorescence; D006801:Humans; D008969:Molecular Sequence Data; D008970:Molecular Weight; D009154:Mutation / Q000235:genetics*; D010455:Peptides / Q000737:chemistry / Q000235:genetics / Q000378:metabolism / Q000494:pharmacology*; D011485:Protein Binding; D011487:Protein Conformation / Q000187:drug effects; D016415:Sequence Alignment; D019065:Virus Assembly / Q000187:drug effects
D003067:Coenzymes; D004220:Disulfides; D010455:Peptides; D003546:Cysteine Endopeptidases; C091827:adenain