Nucleotide sequence and characteristics of the gene for L-lactate dehydrogenase of Thermus aquaticus YT-1 and the deduced amino acid sequence of the enzyme.
The gene for L-lactate dehydrogenase (LDH) from Thermus aquaticus YT-1 was cloned in Escherichia coli, using the Thermus caldophilus LDH gene as a hybridization probe, and its complete nucleotide sequence was determined. The LDH gene comprised 930 base pairs, starting with a GTG initiation codon. Its sequence had high homology (85.8% identity) with the LDH gene of T. caldophilus. The G + C content of the T. aquaticus gene was 70.9%, higher than that of the chromosomal DNA (67.4%). In particular, that in the third position of the codons used was 91.0%, similar to the T. caldophilus gene. The primary structure of T. aquaticus LDH was deduced from the nucleotide sequence of the LDH gene. It comprises 310 amino acid residues, as does T. caldophilus LDH, and its molecular mass was calculated to be 33,210 daltons. The amino acid sequence of the T. aquaticus LDH had 87.1% identity with that of the T. caldophilus LDH. At 23 positions, the respective residues differed in charge and polarity. These differences must be related to the differences in kinetic properties between the two enzymes. The constructed plasmid overproduced the T. aquaticus LDH in E. coli.
Department of Agricultural Chemistry, University of Tokyo.
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Ono
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Matsuzawa
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Ohta
D000595:Amino Acid Sequence; D001483:Base Sequence; D003001:Cloning, Molecular; D003062:Codon; D004269:DNA, Bacterial / Q000235:genetics; D004926:Escherichia coli / Q000235:genetics; D005798:Genes, Bacterial; D007770:L-Lactate Dehydrogenase / Q000235:genetics*; D008969:Molecular Sequence Data; D009693:Nucleic Acid Hybridization; D011994:Recombinant Proteins; D015183:Restriction Mapping; D012689:Sequence Homology, Nucleic Acid; D013824:Thermus / Q000201:enzymology / Q000235:genetics*
D003062:Codon; D004269:DNA, Bacterial; D011994:Recombinant Proteins; D007770:L-Lactate Dehydrogenase