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Lys-34, Dispensable for Integrase Catalysis, Is Required for Preintegration Complex Function and Human Immunodeficiency Virus Type 1 Replication

2005; American Society for Microbiology; Volume: 79; Issue: 19 Linguagem: Inglês

10.1128/jvi.79.19.12584-12591.2005

1098-5514

Richard Lu, Nick Vandegraaff, Peter Cherepanov, Alan Engelman,

Biochemical and Molecular Research

ABSTRACT Retroviral integrases (INs) function in the context of preintegration complexes (PICs). Two conserved Lys residues in the N-terminal domain of human immunodeficiency virus type 1 (HIV-1) IN were analyzed here for their roles in integration and virus replication. Whereas HIV-1 K46A grew like the wild type, HIV-1 K34A was dead. Yet recombinant IN K34A protein functioned in in vitro integration assays, and Vpr-IN K34A efficiently transcomplemented the infectivity defect of an IN active site mutant virus in cells. HIV-1 K34A was therefore similar to a number of previously characterized mutant viruses that failed to replicate despite encoding catalytically competent IN. To directly analyze mutant PIC function, a sensitive PCR-based integration assay was developed. HIV-1 K34A and related mutants failed to support detectable levels (<1% of wild type) of integration. We therefore concluded that mutations like K34A disrupted higher-order interactions important for PIC function/maturation compared to the innate catalytic activity of IN enzyme.