Stabilization of Escherichia coli isopropylmalate dehydrogenase by single amino acid substitution.
To determine the key position for the unusual stability of isopropylmalate dehydrogenase from extreme thermophiles (Thermus thermophilus and T. aquaticus), sequence comparisons were carried out. As a result, a motif which is characteristic to the thermophilic dehydrogenases was found between two highly conserved stretches. The sequence motif was introduced into a mesophilic (Escherichia coli) isopropylmalate dehydrogenase, one by one. Contrary to our expectation, introduction of the whole motif led the mesophilic enzyme to be more unstable whereas substitution of only one amino acid residue in the motif thermostabilized the enzyme. From the 3D structure of the enzyme, a mechanism for the thermostabilization is speculated.
Aoshima
M
M
Department of Molecular Biology, Tokyo University of Pharmacy, Japan.
Oshima
T
T
D050539:3-Isopropylmalate Dehydrogenase; D000429:Alcohol Oxidoreductases / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D000595:Amino Acid Sequence; D001483:Base Sequence; D016208:Databases, Factual; D019281:Dimerization; D004795:Enzyme Stability; D004926:Escherichia coli / Q000201:enzymology*; D006358:Hot Temperature; D008958:Models, Molecular; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D011487:Protein Conformation; D013329:Structure-Activity Relationship
D000429:Alcohol Oxidoreductases; D050539:3-Isopropylmalate Dehydrogenase