Subunit-specific Protein Footprinting Reveals Significant Structural Rearrangements and a Role for N-terminal Lys-14 of HIV-1 Integrase During Viral DNA Binding

Overview

Journal J Biol Chem

Specialty Biochemistry

Date 2007 Dec 21

PMID 18093980

Citations 40

Authors

Zhuojun Zhao

Christopher J McKee

Jacques J Kessl

Webster L Santos

Janet E Daigle

Alan Engelman

Gregory Verdine

Mamuka Kvaratskhelia

Affiliations

Soon will be listed here.

Abstract

To identify functional contacts between HIV-1 integrase (IN) and its viral DNA substrate, we devised a new experimental strategy combining the following two methodologies. First, disulfide-mediated cross-linking was used to site-specifically link select core and C-terminal domain amino acids to respective positions in viral DNA. Next, surface topologies of free IN and IN-DNA complexes were compared using Lys- and Arg-selective small chemical modifiers and mass spectrometric analysis. This approach enabled us to dissect specific contacts made by different monomers within the multimeric complex. The foot-printing studies for the first time revealed the importance of a specific N-terminal domain residue, Lys-14, in viral DNA binding. In addition, a DNA-induced conformational change involving the connection between the core and C-terminal domains was observed. Site-directed mutagenesis experiments confirmed the importance of the identified contacts for recombinant IN activities and virus infection. These new findings provided major constraints, enabling us to identify the viral DNA binding channel in the active full-length IN multimer. The experimental approach described here has general application to mapping interactions within functional nucleoprotein complexes.

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