Kinetic properties and thermal stabilities of mutant forms of mitochondrial aspartate aminotransferase

Amalia Azzariti, Rosa A. Vacca, Sergio Giannattasio, Riccardo S. Merafina, Ersilia Marra, Shawn Doonan

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Kinetic properties and thermal stabilities of the precursor form of mitochondrial aspartate aminotransferase, the mature form lacking 9 amino acids from the N-terminus, and forms of the mature protein in which cysteine-166 had been mutated to serine or alanine were compared with those of the mature enzyme. The precursor and the cysteine mutants showed moderately impaired catalytic properties consistent with decreased ability to undergo transition from the open to the closed conformation which is an integral part of the mechanism of action of the enzyme. The deletion mutant had a k(cat) only 2% of that of the mature enzyme but also much reduced K(m) values for both substrates. In addition it showed enhanced reactivity of cysteine-166 with 5,5'-dithiobis(2-nitrobenzoate), which is characteristic of the closed form of the enzyme, with no enhancement of reactivity in the presence of substrates. This is taken to show that the deletion mutant adopts a conformation that is significantly different from that of the mature enzyme particularly in respect of the small domain. The deletion mutant was found to be more resistant to thermal inactivation over a range of temperatures than were the other forms of the enzyme consistent with its having a more tightly packed small domain. Copyright (C) 1998 Elsevier Science B.V.

Original languageEnglish
Pages (from-to)29-38
Number of pages10
JournalBiochimica et Biophysica Acta (BBA)/Protein Structure and Molecular
Issue number1
Publication statusPublished - Jul 28 1998


  • Aspartate aminotransferase
  • Catalytic property
  • Engineered mutant
  • Thermal stability

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Structural Biology
  • Biophysics


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