The gene encoding glyoxalase I from Pseudomonas putida: cloning, overexpression, and sequence comparisons with human glyoxalase I.
The gene encoding glyoxalase I (GlxI) from Pseudomonas putida has been cloned into the high-expression plasmid pBTacI. In the presence of IPTG, JM109 cells transformed with this vector give expression levels of GlxI 4000-fold higher than wild-type Escherichia coli. Contrary to a previous report, the nucleotide sequence of the gene encodes a 173-amino-acid polypeptide. Edman analysis indicates that the predicted N-terminal methionine is lost post-translationally to yield a 19407-Da protein. Mass spectrometry of the intact protein, and of the peptides generated from treatment with CNBr, does not indicate any additional post-translational modifications of the enzyme. Contrary to previous conclusions, there are no major regions of dissimilarity between the human and bacterial enzymes.
Department of Chemistry, University of Maryland Baltimore County 21228.
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Lovett
D000595:Amino Acid Sequence; D001483:Base Sequence; D003001:Cloning, Molecular; D004269:DNA, Bacterial; D004926:Escherichia coli; D005798:Genes, Bacterial; D006801:Humans; D007791:Lactoylglutathione Lyase / Q000235:genetics*; D013058:Mass Spectrometry; D008969:Molecular Sequence Data; D010957:Plasmids; D016958:Pseudomonas putida / Q000201:enzymology / Q000235:genetics*; D017386:Sequence Homology, Amino Acid
D004269:DNA, Bacterial; D007791:Lactoylglutathione Lyase