Role of histidine 188 in fructose 1,6-bisphosphate- and divalent cation-regulated L-lactate dehydrogenase of Lactobacillus casei.
A fructose 1,6-bisphosphate [Fru(1,6)P2] and divalent cation-regulated allosteric L-lactate dehydrogenase (L-LDH) (EC 1.1.1.27) of Lactobacillus casei was highly produced in Escherichia coli cells, together with its mutant enzyme, in which His-188 was replaced by Asp. Under acidic conditions, the mutant enzyme showed positive allosteric regulations by the substrate pyruvate and its analogues, like the wild-type enzyme, but not by Fru(1,6)P2, which even inhibited the stimulative effects of the alternative activation factors. In addition, Mn2+ ions also showed greatly reduced inhibitory effects on the mutant enzyme. Under neutral conditions, on the other hand, the reaction of the mutant enzyme was slightly enhanced by Fru(1,6)P2, but not further stimulated by additional Mn2+ ions, unlike the case of the wild-type enzyme. These results indicate that His-188 is, though not essential for the regulation by the alternative factors, essential for the cooperative regulation by Fru(1,6)P2 and divalent cations in L. casei L-LDH.
Taguchi
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H
Department of Biotechnology, University of Tokyo, Japan.
Ohta
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T
D000596:Amino Acids / Q000378:metabolism; D001483:Base Sequence; D001665:Binding Sites / Q000502:physiology; D002413:Cations, Divalent / Q000378:metabolism*; D003001:Cloning, Molecular; D004269:DNA, Bacterial / Q000032:analysis / Q000302:isolation & purification; D004926:Escherichia coli / Q000201:enzymology / Q000235:genetics; D005635:Fructosediphosphates / Q000378:metabolism*; D005818:Genetic Engineering; D006639:Histidine / Q000502:physiology*; D006863:Hydrogen-Ion Concentration; D007527:Isoenzymes; D007770:L-Lactate Dehydrogenase / Q000302:isolation & purification / Q000378:metabolism* / Q000502:physiology; D007778:Lactobacillus / Q000201:enzymology / Q000235:genetics; D007780:Lacticaseibacillus casei / Q000201:enzymology*; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed
D000596:Amino Acids; D002413:Cations, Divalent; D004269:DNA, Bacterial; D005635:Fructosediphosphates; D007527:Isoenzymes; D006639:Histidine; D007770:L-Lactate Dehydrogenase; C029063:fructose-1,6-diphosphate