Simian-virus-40 large-T-antigen-catalyzed DNA and RNA unwinding reactions.
Simian virus 40 large T antigen is a helicase separating the complementary strands of double-stranded DNA in the presence of hydrolyzable ATP and of double-stranded RNA in the presence of non-ATP nucleotides (GTP, CTP or UTP). We have constructed partially single-stranded nucleic acid substrates consisting of RNA or DNA strands hydrogen bonded to either RNA or DNA complements. We found that ATP is utilized as a cofactor for the T-antigen-catalyzed unwinding reaction when the substrates contain overhanging single-stranded DNA, regardless of whether the double-stranded region is DNA or hybrid DNA.RNA. Conversely, non-ATP nucleotides are used when the overhanging single strand is RNA. Based on these and additional findings, we propose that the bound nucleic acid induces a conformational change in T antigen resulting in a proper orientation of both nucleic acid and nucleotide relative to the active center of the ATPase/helicase domain of the enzyme. The implications of our conclusion for the roles which T antigen may play in vivo are discussed.
Scheffner
M
M
Fakultät für Biologie, Universität Konstanz, Federal Republic of Germany.
Knippers
R
R
Stahl
H
H
D000911:Antibodies, Monoclonal; D000952:Antigens, Polyomavirus Transforming; D004265:DNA Helicases / Q000378:metabolism*; D004277:DNA, Single-Stranded / Q000378:metabolism; D006860:Hydrogen Bonding; D007700:Kinetics; D009693:Nucleic Acid Hybridization; D020365:RNA Helicases; D012316:RNA Nucleotidyltransferases / Q000378:metabolism*; D012330:RNA, Double-Stranded / Q000378:metabolism*; D012265:Ribonucleotides / Q000494:pharmacology; D013539:Simian virus 40 / Q000201:enzymology / Q000276:immunology*
D000911:Antibodies, Monoclonal; D000952:Antigens, Polyomavirus Transforming; D004277:DNA, Single-Stranded; D012330:RNA, Double-Stranded; D012265:Ribonucleotides; D012316:RNA Nucleotidyltransferases; D004265:DNA Helicases; D020365:RNA Helicases