Adenovirus protein-protein interactions: hexon and protein VI.
A variety of mastadenoviruses were denatured, their polypeptides separated by electrophoresis on SDS-polyacrylamide gels and transferred to nitrocellulose. The immobilized polypeptides were washed, incubated with buffers containing hexons from human adenoviruses (Ad) types 2, 5 and 12 and the location of bound hexons was detected with anti-hexon antibodies. It was found that hexons from any of the three human adenovirus types bound to protein VI from all the mastadenoviruses examined. Furthermore we found that hexon-VI binding was significantly greater than the interaction between hexon and the precursor to VI, pVI. This binding was susceptible to detergents and to changes in pH or salt concentration. A rabbit polyclonal antibody was raised against a recombinant protein derived from the middle third of pVI from Ad2 and was used to quantify the difference in binding and to demonstrate the presence of a single intermediate (designated iVI) in the processing of pVI to VI. The affinity between iVI and hexon was considerably greater in our assay than that of pVI but was less than that between hexon and VI. A complementary binding of recombinant iVI to immobilized hexons was also demonstrated. This latter interaction, however, was only observed when hexon preparations were not boiled prior to electrophoresis, substantiating the proposition that the recognition motif on the hexon was conformation-dependent. These results are discussed in the context of understanding further the molecular basis of protein-protein interactions between the structural proteins of adenoviruses and the factors involved in virion maturation.
Matthews
D A
DA
Division of Cell and Molecular Biology, School of Biological and Medical Sciences, University of St. Andrews, Fife, U.K.
Russell
W C
WC
D000260:Adenoviruses, Human / Q000737:chemistry*; D000595:Amino Acid Sequence; D000914:Antibodies, Viral; D001483:Base Sequence; D015153:Blotting, Western / Q000379:methods; D002213:Capsid / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D036022:Capsid Proteins; D003001:Cloning, Molecular; D004926:Escherichia coli / Q000235:genetics; D015870:Gene Expression; D005814:Genes, Viral / Q000235:genetics; D006801:Humans; D017921:Mastadenovirus / Q000737:chemistry*; D008969:Molecular Sequence Data; D008970:Molecular Weight; D011489:Protein Denaturation; D011498:Protein Precursors / Q000378:metabolism; D011994:Recombinant Proteins / Q000302:isolation & purification / Q000378:metabolism; D017422:Sequence Analysis, DNA; D013045:Species Specificity; D015678:Viral Structural Proteins / Q000235:genetics
D000914:Antibodies, Viral; D036022:Capsid Proteins; D011498:Protein Precursors; D011994:Recombinant Proteins; D015678:Viral Structural Proteins; C018315:hexon capsid protein, Adenovirus; C090876:polypeptide VI, adenovirus