A peptide fragment of human DNA topoisomerase II alpha forms a stable coiled-coil structure in solution.
Results are presented on a peptide fragment (1013-1056) from human DNA topoisomerase II alpha. This was selected using the procedure of Lupas et al. (Lupas, A., Van Dyke, M., and Stock, J. (1991) Science 252, 1162-1164) for its potential to adopt a stable coiled-coil structure. The same theoretical treatment rejected the segment 994-1021 proposed by Zwelling and Perry (Zwelling, L. A., and Perry, W. M. (1989) Mol. Endocrinol. 3, 603-604) as a possible core for leucine-zipper formation. Our experimental studies combine cross-linking and CD analysis. Cross-linking establishes that the 1013-1056 fragment forms a stable homodimer in solution. Effects of increasing peptide concentration on CD spectra confirm that only the 1013-1056 fragment can undergo a coiled-coil stabilization from an isolated alpha-helix. Unfolding experiments further show that the coiled-coil is more stable in guanidium chloride than in urea. Values of -6.8 and -7.4 kcal/mol for the dimerization free energy are determined by thermal and urea unfolding, respectively. These are strikingly similar to the value recently found for the dissociation/reassociation of the entire yeast topoisomerase II from sedimentation equilibrium experiments (Lamhasni, S., Larsen, A. K., Barray, M., Monnot, M., Delain, E., and Fermandjian, S. (1995) Biochemistry 34, 3632-3639), although their significance relatively to topoisomerase II undoubtedly requires further analysis.
Frère
V
V
Département de Biologie et Pharmacologie Structurales, URA 147 CNRS, Institut Gustave Roussy, Villejuif, France.
Sourgen
F
F
Monnot
M
M
Troalen
F
F
Fermandjian
S
S
D000595:Amino Acid Sequence; D002942:Circular Dichroism; D004250:DNA Topoisomerases, Type II / Q000737:chemistry*; D005976:Glutaral / Q000494:pharmacology; D019791:Guanidine; D006146:Guanidines / Q000494:pharmacology; D006801:Humans; D008969:Molecular Sequence Data; D010446:Peptide Fragments / Q000737:chemistry*; D017510:Protein Folding; D012996:Solutions; D014270:Trifluoroethanol / Q000494:pharmacology; D014508:Urea / Q000494:pharmacology
D006146:Guanidines; D010446:Peptide Fragments; D012996:Solutions; D014270:Trifluoroethanol; D014508:Urea; D004250:DNA Topoisomerases, Type II; D019791:Guanidine; D005976:Glutaral