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Oligomeric integrity—The structural key to thermal stability in bacterial alcohol dehydrogenases

Published online by Cambridge University Press:  01 June 1999

YAKOV KORKHIN ,
A. JOSEPH KALB(GILBOA) ,
MOSHE PERETZ ,
OREN BOGIN ,
YIGAL BURSTEIN  and
FELIX FROLOW
YAKOV KORKHIN
Affiliation:
Department of Structural Biology, The Weizmann Institute of Science, Rehovot 76100, Israel Present address: Department of Molecular Biophysics and Biochemistry, Yale University, New Haven, Connecticut 06520-8114.
A. JOSEPH KALB(GILBOA)
Affiliation:
Department of Structural Biology, The Weizmann Institute of Science, Rehovot 76100, Israel
MOSHE PERETZ
Affiliation:
Department of Organic Chemistry, The Weizmann Institute of Science, Rehovot 76100, Israel
OREN BOGIN
Affiliation:
Department of Organic Chemistry, The Weizmann Institute of Science, Rehovot 76100, Israel
YIGAL BURSTEIN
Affiliation:
Department of Organic Chemistry, The Weizmann Institute of Science, Rehovot 76100, Israel
FELIX FROLOW
Affiliation:
Department of Chemical Services, The Weizmann Institute of Science, Rehovot 76100, Israel Present address: Department of Molecular Microbiology and Biotechnology, The George S. Wise Faculty of Life Sciences, Tel-Aviv University, Ramat Aviv 69978, Israel.
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Abstract

Principles of protein thermostability have been studied by comparing structures of thermostable proteins with mesophilic counterparts that have a high degree of sequence identity. Two tetrameric NADP(H)-dependent alcohol dehydrogenases, one from Clostridium beijerinckii (CBADH) and the other from Thermoanaerobacter brockii (TBADH), having exceptionally high (75%) sequence identity, differ by 30° in their melting temperatures. The crystal structures of CBADH and TBADH in their holo-enzyme form have been determined at a resolution of 2.05 and 2.5 Å, respectively. Comparison of these two very similar structures (RMS difference in Cα = 0.8 Å) revealed several features that can account for the higher thermal stability of TBADH. These include additional ion pairs, “charged-neutral” hydrogen bonds, and prolines as well as improved stability of α-helices and tighter molecular packing. However, a deeper structural insight, based on the location of stabilizing elements, suggests that enhanced thermal stability of TBADH is due mainly to the strategic placement of structural determinants at positions that strengthen the interface between its subunits. This is also supported by mutational analysis of structural elements at critical locations. Thus, it is the reinforcement of the quaternary structure that is most likely to be a primary factor in preserving enzymatic activity of this oligomeric bacterial ADH at elevated temperatures.

Keywords

alcohol dehydrogenasecrystal structureoligomersmutagenesisthermal stability
Type
Research Article
Information
Protein Science , Volume 8 , Issue 6 , June 1999 , pp. 1241 - 1249
Copyright
© 1999 The Protein Society

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