Structural implications of spectroscopic characterization of a putative zinc finger peptide from HIV-1 integrase.

The solution structure of the amino-terminal HHCC domain of HIV-2 integrase: a three-helix bundle stabilized by zinc

“…IN 1-55 was further purified by gel filtration on a HiLoad Superdex 75 column (Pharmacia) and concentrated using Centriprep (Amicon). The protein was dialysed to 3 M guanidine-HCl, 20 mM Tris, pH 8.3, 3 mM β-mercaptoethanol, followed by a reduction step in 20% (v/v) β-mercaptoethanol at 55°C for 1.5 h as described by Burke et al [20]. Subsequently, the protein was purified over a C-18 reversed phase HPLC column and eluted with a gradient of 5-70% (v/v) acetonitrile in 0.05% (v/v) trifluoroacetic acid.…”

“…Zinc binding by the first 55 amino acids of integrase has been demonstrated in a zinc blot and by atomic absorption spectroscopy [6,20,21]. Zinc induces folding in the aminoterminal domain as well as in the full-length protein and binds in an equimolar ratio [20,22]. In the full-length protein, the presence of zinc promotes multimerization and stimulates Mg 2+ -dependent cleavage [22][23][24].…”

“…Although the exact role of this domain is not known, it is required for cleavage and integration. Zinc binding by the first 55 amino acids of integrase has been demonstrated in a zinc blot and by atomic absorption spectroscopy [6,20,21]. Zinc induces folding in the aminoterminal domain as well as in the full-length protein and binds in an equimolar ratio [20,22].…”

“…The primary sequence of the amino-terminal domain does not show homology with any of the zinc-finger topologies characterized thus far [27][28][29][30]. Previously, it has been suggested that the structure of this domain might have similarities with the structure of the CCHH zinc-finger domain, which is found in transcription factor IIIA-like peptides [20]. However, the three-dimensional solution structure of the amino-terminal domain of HIV-2 integrase, as reported here, shows no structural similarity with any of the known zinc-finger structures.…”

Multivalent interactions essential for lentiviral integrase function

“…The side chains of Arg231 residues from various IN chains are directly involved in interactions with viral and target DNA (Figs 1c and 2a). Finally, MVV IN His12 is part of the invariant HHCC motif that coordinates a Zn 2+ ion, which is critical for NTD structural stability (39, 40). Within the MVV intasome, the NTDs from numerous IN chains contribute to the CIC and/or to IN-vDNA and IN-IN interactions (Figs 1a, 1d, and S3b).…”