Loop202-208 in avian sarcoma virus integrase mediates tetramer assembly and processing activity.
Integrase (IN) catalyzes insertion of the retroviral genome into the host via two sequential reactions. The processing activity cleaves the 3'-dinucleotides from the two ends of the viral DNA which are then inserted into the host DNA. Tetramers are required for the joining step. While dimers have been shown to catalyze processing, they do so inefficiently, and the oligomeric requirement for processing is unknown. We have replaced loop202-208 at the putative dimer-dimer interface of the avian sarcoma virus IN with its analogue, loop188-194, from human immunodeficiency virus IN. The mutation abolished disintegration activity and a 2 x 10(-2) s-1 fast phase during single-turnover processing. A 3 x 10(-4) s-1 slow processing phase was unaffected. Preincubation with a DNA substrate known to promote tetramerization increased products formed during the fast phase by 2.5-fold only for wild-type IN, correlating the fast and slow phases with processing by tetramers and dimers, respectively. We propose a novel tetramer model for coupling processing and integration based on efficient processing by the tetramer. We provide for the first time direct evidence of the functional relevance of a structural element, loop202-208, which appears to be required for mediating the interaction between dimer halves of the active tetramer.
Bosserman
Mary A
MA
Department of Molecular and Cellular Biochemistry, University of Kentucky, Lexington, Kentucky 40536, USA.
O'Quinn
Daniel F
DF
Wong
Isaac
I
D000595:Amino Acid Sequence; D001358:Avian Sarcoma Viruses / Q000737:chemistry / Q000201:enzymology* / Q000235:genetics; D001483:Base Sequence; D019281:Dimerization; D019427:HIV Integrase / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D019426:Integrases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D007700:Kinetics; D008958:Models, Molecular; D008969:Molecular Sequence Data; D016296:Mutagenesis; D017433:Protein Structure, Secondary; D013329:Structure-Activity Relationship; D014764:Viral Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism
D014764:Viral Proteins; D019427:HIV Integrase; D019426:Integrases