Mirror image mutations reveal the significance of an intersubunit ion cluster in the stability of 3-isopropylmalate dehydrogenase

Attila Németh, Ádám Svingor, Márta Pócsik, József Dobó, Csaba Magyar, András Szilágyi, Péter Gál, Péter Závodszky

Research output: Contribution to journalArticle

15 Citations (Scopus)

Abstract

The comparison of the three-dimensional structures of thermophilic (Thermus thermophilus) and mesophilic (Escherichia coli) 3-isopropylmalate dehydrogenases (IPMDH, EC 1.1.1.85) suggested that the existence of extra ion pairs in the thermophilic enzyme found in the intersubunit region may be an important factor for thermostability. As a test of our assumption, glutamine 200 in the E. coli enzyme was turned into glutamate (Q200E mutant) to mimic the thermophilic enzyme at this site by creating an intersubunit ion pair which can join existing ion clusters. At the same site in the thermophilic enzyme we changed glutamate 190 into glutamine (E190Q), hereby removing the corresponding ion pair. These single amino acid replacements resulted in increased thermostability of the mesophilic and decreased thermostability of the thermophilic enzyme, as measured by spectropolarimetry and differential scanning microcalorimetry. Copyright (C) 2000 Federation of European Biochemical Societies.

Original languageEnglish
Pages (from-to)48-52
Number of pages5
JournalFEBS letters
Volume468
Issue number1
DOIs
Publication statusPublished - Feb 18 2000

Keywords

  • 3-Isopropylmalate dehydrogenase
  • Circular dichroism
  • Differential scanning calorimetry
  • Ion cluster
  • Site-directed mutagenesis
  • Thermostability

ASJC Scopus subject areas

  • Biophysics
  • Structural Biology
  • Biochemistry
  • Molecular Biology
  • Genetics
  • Cell Biology

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