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Three new structures of the core domain of HIV-1 integrase: an active site that binds magnesium.

ABSTRACT: HIV-1 integrase is an essential enzyme in the life cycle of the virus, responsible for catalyzing the insertion of the viral genome into the host cell chromosome; it provides an attractive target for antiviral drug design. The previously reported crystal structure of the HIV-1 integrase core domain revealed that this domain belongs to the superfamily of polynucleotidyltransferases. However, the position of the conserved catalytic carboxylic acids differed from those observed in other enzymes of the class, and attempts to crystallize in the presence of the cofactor, Mg2+, were unsuccessful. We report here three additional crystal structures of the core domain of HIV-1 integrase mutants, crystallized in the presence and absence of cacodylate, as well as complexed with Mg2+. These three crystal forms, containing between them seven independent core domain structures, demonstrate the unambiguous extension of the previously disordered helix alpha4 toward the amino terminus from residue M154 and show that the catalytic E152 points in the general direction of the two catalytic aspartates, D64 and D116. In the vicinity of the active site, the structure of the protein in the absence of cacodylate exhibits significant deviations from the previously reported structures. These differences can be attributed to the modification of C65 and C130 by cacodylate, which was an essential component of the original crystallization mixture. We also demonstrate that in the absence of cacodylate this protein will bind to Mg2+, and could provide a satisfactory platform for binding of inhibitors.

SUBMITTER: Goldgur Y

PROVIDER: S-EPMC21307 | biostudies-literature | 1998 Aug

REPOSITORIES: biostudies-literature

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Three new structures of the core domain of HIV-1 integrase: an active site that binds magnesium.

Goldgur Y Y Dyda F F Hickman A B AB Jenkins T M TM Craigie R R Davies D R DR

Proceedings of the National Academy of Sciences of the United States of America 19980801 16

HIV-1 integrase is an essential enzyme in the life cycle of the virus, responsible for catalyzing the insertion of the viral genome into the host cell chromosome; it provides an attractive target for antiviral drug design. The previously reported crystal structure of the HIV-1 integrase core domain revealed that this domain belongs to the superfamily of polynucleotidyltransferases. However, the position of the conserved catalytic carboxylic acids differed from those observed in other enzymes of ...[more]

PMID: 9689049

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Project description:HIV integrase (HIV-IN), one of three HIV enzymes, is a target for the treatment of AIDS, but the full biological assembly has been difficult to characterize, hampering inhibitor design. The recent crystallographic structures of integrase from prototype foamy virus (PFV-IN) with bound DNA were a breakthrough, revealing how viral DNA organizes two integrase dimers into a tetramer that has the two active sites appropriately spaced for insertion of the viral DNA into host DNA. The organization of domains within each PFV-IN protein chain, however, varies significantly from that found in HIV-IN structures. With the goal of identifying shared structural characteristics, the interactions among components of the PFV-IN and HIV-IN assemblies were investigated with the macromolecular docking program DOT. DOT performs an exhaustive, rigid-body search between two macromolecules. Computational docking reproduced the crystallographic interactions of the PFV-IN catalytic and N-terminal domains with viral DNA and found similar viral DNA interactions for HIV-IN. Computational docking did not reproduce the crystallographic interactions of the PFV-IN C-terminal domain (CTD). Instead, two symmetry-related positions were found for the PFV-IN CTD that indicate formation of a CTD dimer between the two active sites. Our predicted CTD dimer is consistent with cross-linking studies showing interactions of the CTD with viral DNA that appear to be blocked in the PFV-IN structures. The CTD dimer can insert two arginine-rich loops between the two bound vDNA molecules and the host DNA, a region that is unoccupied in the PFV-IN crystallographic structures. The positive potential from these two loops would alleviate the large negative potential created by the close proximity of two viral vDNA ends, helping to bring together the two active sites and assisting host DNA binding. This study demonstrates the ability of computational docking to evaluate complex crystallographic assemblies, identify interactions that are influenced by the crystal environment, and provide plausible alternatives.

Dataset Information

Three new structures of the core domain of HIV-1 integrase: an active site that binds magnesium.

Publications

Three new structures of the core domain of HIV-1 integrase: an active site that binds magnesium.

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