The initial step of the thermal unfolding of 3-isopropylmalate dehydrogenase detected by the temperature-jump Laue method

Tetsuya Hori, Hideaki Moriyama, Jitsutaro Kawaguchi, Yoko Hayashi-Iwasaki, Tairo Oshima, Nobuo Tanaka

Research output: Contribution to journalArticle

15 Citations (Scopus)

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 (~90°C) was found to be localized at a region which includes a β-turn and a loop located between the two domains of the enzyme. A mutant, 2T2M6T-E110P/S111G/S113E, having amino acid replacements in this β-turn region with the corresponding residues of the thermophilic enzyme, showed greater stability than the original chimera (increase of T(m) by ~10°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 β-turn region.

Original languageEnglish (US)
Pages (from-to)527-533
Number of pages7
JournalProtein Engineering
Volume13
Issue number8
StatePublished - Oct 3 2000

Fingerprint

3-Isopropylmalate Dehydrogenase
Hot Temperature
Temperature
Enzymes
Proteins
Laser beam effects
Amino acids
Lasers
X-Rays
Oxidoreductases
Amino Acids
X rays
Crystals

Keywords

  • 3-Isopropylmalate dehydrogenase
  • Crystal structure
  • Temperature-jump Laue method
  • Thermal unfolding
  • Thermostability

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology

Cite this

The initial step of the thermal unfolding of 3-isopropylmalate dehydrogenase detected by the temperature-jump Laue method. / Hori, Tetsuya; Moriyama, Hideaki; Kawaguchi, Jitsutaro; Hayashi-Iwasaki, Yoko; Oshima, Tairo; Tanaka, Nobuo.

In: Protein Engineering, Vol. 13, No. 8, 03.10.2000, p. 527-533.

Research output: Contribution to journalArticle

Hori, T, Moriyama, H, Kawaguchi, J, Hayashi-Iwasaki, Y, Oshima, T & Tanaka, N 2000, 'The initial step of the thermal unfolding of 3-isopropylmalate dehydrogenase detected by the temperature-jump Laue method', Protein Engineering, vol. 13, no. 8, pp. 527-533.
Hori, Tetsuya ; Moriyama, Hideaki ; Kawaguchi, Jitsutaro ; Hayashi-Iwasaki, Yoko ; Oshima, Tairo ; Tanaka, Nobuo. / The initial step of the thermal unfolding of 3-isopropylmalate dehydrogenase detected by the temperature-jump Laue method. In: Protein Engineering. 2000 ; Vol. 13, No. 8. pp. 527-533.
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