title: The initial step of the thermal unfolding of 3-isopropylmalate dehydrogenase detected by the temperature-jump Laue method.
abstract: A temperature-jump (T-jump) time-resolved X-ray crystallographic technique using the Laue method was developed to detect small, localized structural changes of proteins in crystals exposed to a temperature increase induced by laser irradiation. In a chimeric protein between thermophilic and mesophilic 3-isopropylmalate dehydrogenases (2T2M6T), the initial structural change upon T-jump to a denaturing temperature (approximately 90 degrees C) was found to be localized at a region which includes a beta-turn and a loop located between the two domains of the enzyme. A mutant, 2T2M6T-E110P/S111G/S113E, having amino acid replacements in this beta-turn region with the corresponding residues of the thermophilic enzyme, showed greater stability than the original chimera (increase of T:(m) by approximately 10 degrees C) and no T-jump-induced structural change in this region was detected by our method. These results indicate that thermal unfolding of the original chimeric enzyme, 2T2M6T, is triggered in this beta-turn region.
authors.affiliation: Graduate School of Bioscience and Biotechnology, Tokyo Institute of Technology, Nagatsuta 4259, Midori-ku, Yokohama 226-8501, Japan.
authors.forename: T
authors.initials: T
authors.lastname: Hori
authors.forename: H
authors.initials: H
authors.lastname: Moriyama
authors.forename: J
authors.initials: J
authors.lastname: Kawaguchi
authors.forename: Y
authors.initials: Y
authors.lastname: Hayashi-Iwasaki
authors.forename: T
authors.initials: T
authors.lastname: Oshima
authors.forename: N
authors.initials: N
authors.lastname: Tanaka
mesh_terms: D050539:3-Isopropylmalate Dehydrogenase; D000429:Alcohol Oxidoreductases / Q000737:chemistry*; D000595:Amino Acid Sequence; D002942:Circular Dichroism; D008969:Molecular Sequence Data; D011487:Protein Conformation; D011489:Protein Denaturation; D013696:Temperature; D014961:X-Ray Diffraction
chemical_list: D000429:Alcohol Oxidoreductases; D050539:3-Isopropylmalate Dehydrogenase