Format

Send to

Choose Destination
Biochemistry. 2001 Oct 16;40(41):12349-56.

Interaction of the human adenovirus proteinase with its 11-amino acid cofactor pVIc.

Author information

1
Department of Pharmacological Sciences, State University of New York at Stony Brook, Stony Brook, New York 11794, USA.

Erratum in

  • Biochemistry 2002 Jan 8;41(1):430.

Abstract

The interaction of the human adenovirus proteinase (AVP) and AVP-DNA complexes with the 11-amino acid cofactor pVIc was characterized. The equilibrium dissociation constant for the binding of pVIc to AVP was 4.4 microM. The binding of AVP to 12-mer single-stranded DNA decreased the K(d) for the binding of pVIc to AVP to 0.09 microM. The pVIc-AVP complex hydrolyzed the substrate with a Michaelis constant (K(m)) of 3.7 microM and a catalytic rate constant (k(cat)) of 1.1 s(-1). In the presence of DNA, the K(m) increased less than 2-fold, and the k(cat) increased 3-fold. Alanine-scanning mutagenesis was performed to determine the contribution of individual pVIc side chains in the binding and stimulation of AVP. Two amino acid residues, Gly1' and Phe11', were the major determinants in the binding of pVIc to AVP, while Val2' and Phe11' were the major determinants in stimulating enzyme activity. Binding of AVP to DNA greatly suppressed the effects of the alanine substitutions on the binding of mutant pVIcs to AVP. Binding of either or both of the cofactors, pVIc or the viral DNA, to AVP did not dramatically alter its secondary structure as determined by vacuum ultraviolet circular dichroism. pVIc, when added to Hep-2 cells infected with adenovirus serotype 5, inhibited the synthesis of infectious virus, presumably by prematurely activating the proteinase so that it cleaved virion precursor proteins before virion assembly, thereby aborting the infection.

PMID:
11591154
PMCID:
PMC3590020
DOI:
10.1021/bi0109008
[Indexed for MEDLINE]
Free PMC Article

Supplemental Content

Full text links

Icon for American Chemical Society Icon for PubMed Central
Loading ...
Support Center