A family of highly conserved glycosomal 2-hydroxyacid dehydrogenases from Phytomonas sp.
Phytomonas sp. contains two malate dehydrogenase isoforms, a mitochondrial isoenzyme with a high specificity for oxaloacetate and a glycosomal isozyme that acts on a broad range of substrates (Uttaro, A. D., and Opperdoes, F.R. (1997) Mol. Biochem. Parasitol. 89, 51-59). Here, we show that the low specificity of the latter isoenzyme is the result of a number of recent gene duplications that gave rise to a family of glycosomal 2-hydroxyacid dehydrogenase genes. Two of these genes were cloned, sequenced, and overexpressed in Escherichia coli. Although both gene products have 322 amino acids, share 90.4% identical residues, and have a similar hydrophobicity profile and net charge, their kinetic properties were strikingly different. One isoform behaved as a real malate dehydrogenase with a high specificity for oxaloacetate, whereas the other showed no activity with oxaloacetate but was able to reduce other oxoacids, such as phenyl pyruvate, 2-oxoisocaproate, 2-oxovalerate, 2-oxobutyrate, 2-oxo-4-methiolbutyrate, and pyruvate.
Uttaro
A D
AD
Departamento de MicrobiologIa, Facultad de Ciencias Bioquimicas y Farmacéuticas, Universidad Nacional de Rosario, Suipacha 531, 2000 Rosario, Santa Fe, Argentina.
Altabe
S G
SG
Rider
M H
MH
Michels
P A
PA
Opperdoes
F R
FR
D000429:Alcohol Oxidoreductases / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D000595:Amino Acid Sequence; D000818:Animals; D015139:Blotting, Southern; D003001:Cloning, Molecular; D017124:Conserved Sequence / Q000235:genetics*; D018628:Gene Dosage; D020131:Genes, Duplicate / Q000235:genetics; D017125:Genes, Protozoan / Q000235:genetics; D007527:Isoenzymes / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D007700:Kinetics; D008291:Malate Dehydrogenase / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D008830:Microbodies / Q000201:enzymology*; D008969:Molecular Sequence Data; D005810:Multigene Family / Q000235:genetics*; D010071:Oxaloacetates / Q000378:metabolism; D010802:Phylogeny; D011994:Recombinant Proteins / Q000378:metabolism; D016415:Sequence Alignment; D017422:Sequence Analysis, DNA; D013379:Substrate Specificity; D014351:Trypanosomatina / Q000201:enzymology* / Q000235:genetics
D007527:Isoenzymes; D010071:Oxaloacetates; D011994:Recombinant Proteins; D000429:Alcohol Oxidoreductases; C083779:2-hydroxyacid dehydrogenase; D008291:Malate Dehydrogenase