The role of lysine 186 in HIV-1 integrase multimerization.
HIV-1 integrase (IN) catalyzes biochemical reactions required for viral cDNA insertion into host cell chromosomal DNA, an essential step in the HIV-1 replication cycle. In one of these reactions, the two ends of the linear viral cDNA are believed to be simultaneously ligated to chromosomal DNA by a tetrameric form of IN. The structure of the full-length IN tetramer is not known but a model consisting of the N-terminal domain and the catalytic core revealed basic residues 186 to 188 at the interface between the two IN dimers. We found that alteration of these residues, in particular changing IN lysine residue 186 to glutamate (K186Q), impairs IN oligomerization in the yeast two-hybrid system and decreases oligomeric forms of IN within virions. When expressed independently of other viral proteins in human cells, IN-K186Q did not concentrate in the nucleus as did wild-type IN. Co-expression of wild-type IN restored the multimerization defects of IN-K186Q, in both the two-hybrid system and in virions, and also rescued the nuclear targeting defects. Virions bearing IN-K186Q were not infectious in a single cycle of replication but when mixed virions containing two different IN mutants were produced, IN-K186Q was capable of complementing the catalytically inactive mutant IN-D116A. Our biochemical and functional data support the crystallographic model in which IN residue K186 lies at the interface between IN dimers and suggest that tetramerization is important, not only for concerted integration, but also for IN nuclear targeting.
Berthoux
Lionel
L
Department of Microbiology, Columbia University, New York, USA.
Sebastian
Sarah
S
Muesing
Mark A
MA
Luban
Jeremy
J
D019943:Amino Acid Substitution; D001483:Base Sequence; D002460:Cell Line; D004279:DNA, Viral / Q000235:genetics; D019427:HIV Integrase / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D015497:HIV-1 / Q000201:enzymology* / Q000235:genetics / Q000472:pathogenicity / Q000502:physiology; D006367:HeLa Cells; D006801:Humans; D066298:In Vitro Techniques; D019169:Jurkat Cells; D008239:Lysine / Q000737:chemistry; D008958:Models, Molecular; D016297:Mutagenesis, Site-Directed; D020836:Protein Structure, Quaternary; D020798:Two-Hybrid System Techniques; D014779:Virus Replication
D004279:DNA, Viral; D019427:HIV Integrase; D008239:Lysine