Solution conformation and dynamics of the HIV-1 integrase core domain.
The human immunodeficiency virus type 1 (HIV-1) integrase (IN) is a critical enzyme involved in infection. It catalyzes two reactions to integrate the viral cDNA into the host genome, 3' processing and strand transfer, but the dynamic behavior of the active site during catalysis of these two processes remains poorly characterized. NMR spectroscopy can reveal important structural details about enzyme mechanisms, but to date the IN catalytic core domain has proven resistant to such an analysis. Here, we present the first NMR studies of a soluble variant of the catalytic core domain. The NMR chemical shifts are found to corroborate structures observed in crystals, and confirm prior studies suggesting that the alpha4 helix extends toward the active site. We also observe a dramatic improvement in NMR spectra with increasing MgCl(2) concentration. This improvement suggests a structural transition not only near the active site residues but also throughout the entire molecule as IN binds Mg(2+). In particular, the stability of the core domain is linked to the conformation of its C-terminal helix, which has implications for relative domain orientation in the full-length enzyme. (15)N relaxation experiments further show that, although conformationally flexible, the catalytic loop of IN is not fully disordered in the absence of DNA. Indeed, automated chemical shift-based modeling of the active site loop reveals several stable clusters that show striking similarity to a recent crystal structure of prototype foamy virus IN bound to DNA.
Fitzkee
Nicholas C
NC
Laboratory of Chemical Physics, NIDDK, National Institutes of Health, Bethesda, Maryland 20892, USA.
Masse
James E
JE
Shen
Yang
Y
Davies
David R
DR
Bax
Ad
A
D020134:Catalytic Domain; D018076:DNA, Complementary / Q000378:metabolism; D019427:HIV Integrase / Q000737:chemistry*; D015497:HIV-1 / Q000201:enzymology*; D007700:Kinetics; D008274:Magnesium / Q000737:chemistry; D015636:Magnesium Chloride / Q000737:chemistry; D009682:Magnetic Resonance Spectroscopy; D008345:Manganese / Q000737:chemistry; D008958:Models, Molecular; D008968:Molecular Conformation; D011485:Protein Binding; D011487:Protein Conformation; D017434:Protein Structure, Tertiary; D011760:Pyrrolidinones / Q000737:chemistry; D000068898:Raltegravir Potassium
D018076:DNA, Complementary; D011760:Pyrrolidinones; D015636:Magnesium Chloride; D008345:Manganese; D000068898:Raltegravir Potassium; D019427:HIV Integrase; D008274:Magnesium