Unusual amino acid substitution in the anion-binding site of Lactobacillus plantarum non-allosteric L-lactate dehydrogenase.
In Lactobacillus plantarum non-allosteric L-lactate dehydrogenase (L-LDH), the highly conserved His188 residue, which is involved in the binding of an allosteric effector, fructose 1,6-bisphosphate [Fru(1,6)P2], in allosteric L-LDH is uniquely substituted by an Asp. The mutant L. plantarum L-LDH, in which Asp188 is replaced by a His, showed essentially the same Fru(1,6)P2-independent catalytic activity as the wild-type enzyme, except that the Km and Vmax values were slightly decreased. However, the addition of Fru(1,6)P2 induced significant thermostabilization of the mutant enzyme, as in the case of many allosteric L-LDHs, while Fru(1,6)P2 showed no significant effect on the stability of the wild-type enzyme, indicating that only the single-point mutation, G-->C, sufficiently induces the Fru(1,6)P2-binding ability of L. plantarum L-LDH. The mutant enzyme showed higher thermostability than the wild-type enzyme in the presence of Fru(1,6)P2. In the absence of Fru(1,6)P2, on the other hand, the mutant enzyme was more labile below 65 degrees C but more stable above 70 degrees C.
Taguchi
H
H
Department of Agricultural Chemistry, University of Tokyo, Japan.
Ohta
T
T
D000494:Allosteric Regulation; D000595:Amino Acid Sequence; D000596:Amino Acids / Q000235:genetics*; D000838:Anions; D001483:Base Sequence; D001665:Binding Sites; D002384:Catalysis; D003001:Cloning, Molecular; D004269:DNA, Bacterial; D004795:Enzyme Stability; D004926:Escherichia coli; D005635:Fructosediphosphates / Q000378:metabolism; D006358:Hot Temperature; D006863:Hydrogen-Ion Concentration; D007770:L-Lactate Dehydrogenase / Q000235:genetics* / Q000378:metabolism; D007778:Lactobacillus / Q000201:enzymology*; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D017386:Sequence Homology, Amino Acid
D000596:Amino Acids; D000838:Anions; D004269:DNA, Bacterial; D005635:Fructosediphosphates; D007770:L-Lactate Dehydrogenase; C029063:fructose-1,6-diphosphate