Biophysical and enzymatic properties of the catalytic domain of HIV-1 integrase.
A deletion derivative of the integrase protein from human immunodeficiency virus type-1 (HIV-1) consisting of the central core domain (amino acids 50-212) has been characterized biophysically and biochemically. This deletion mutant is of particular interest for structural studies as it can carry out the disintegration reaction suggesting the presence of an active site and, under certain conditions, is more soluble than full-length integrase. The circular dichroism and fluorescence of the deletion mutant and the 288-residue full-length integrase were similar, indicating that the core residues maintain similar overall conformations in both proteins. The deletion mutant is approximately 10% more alpha-helical than the full-length protein. Analytical centrifugation demonstrated that both proteins undergo monomer-dimer association although the truncated protein showed slightly less tendency to dimerize; the dissociation constants were 2.5 x 10(-5) M for the full-length protein and 8.0 x 10(-5) M for the truncated protein. The disintegration activity of both proteins was also compared. Although a higher concentration of the truncation mutant was required for optimal activity, the mutant did not have altered pH or Mn2+ requirements relative to the full-length protein. The combined biophysical and enzymatic studies suggest that this truncated form of HIV-1 integrase is likely to be useful for structural studies.
Laboratory of Molecular Biology, NIDDK, National Institutes of Health, Bethesda, Maryland 20892.
A B
AB
Hickman
I
I
Palmer
A
A
Engelman
R
R
Craigie
P
P
Wingfield
D000595:Amino Acid Sequence; D001665:Binding Sites; D002384:Catalysis; D002846:Chromatography, Affinity; D002850:Chromatography, Gel; D002942:Circular Dichroism; D004254:DNA Nucleotidyltransferases / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism*; D004591:Electrophoresis, Polyacrylamide Gel; D015497:HIV-1 / Q000201:enzymology*; D019426:Integrases; D008969:Molecular Sequence Data; D008970:Molecular Weight
D004254:DNA Nucleotidyltransferases; D019426:Integrases