Identification, expression, and characterization of Escherichia coli guanine deaminase.
Using the human cDNA sequence corresponding to guanine deaminase, the Escherichia coli genome was scanned using the Basic Local Alignment Search Tool (BLAST), and a corresponding 439-residue open reading frame of unknown function was identified as having 36% identity to the human protein. The putative gene was amplified, subcloned into the pMAL-c2 vector, expressed, purified, and characterized enzymatically. The 50.2-kDa protein catalyzed the conversion of guanine to xanthine, having a K(m) of 15 microM with guanine and a k(cat) of 3.2 s(-1). The bacterial enzyme shares a nine-residue heavy metal binding site with human guanine deaminase, PG[FL]VDTHIH, and was found to contain approximately 1 mol of zinc per mol of subunit of protein. The E. coli guanine deaminase locus is 3' from an open reading frame which shows homology to a bacterial purine base permease.
Departments of Medical Genetics and Biochemistry & Molecular Biology, University of Calgary, Calgary, Alberta T2N 4N1, Canada.
J T
JT
Maynes
R G
RG
Yuan
F F
FF
Snyder
D000595:Amino Acid Sequence; D003001:Cloning, Molecular; D018076:DNA, Complementary; D004926:Escherichia coli / Q000201:enzymology* / Q000235:genetics; D006148:Guanine Deaminase / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D006801:Humans; D007700:Kinetics; D008969:Molecular Sequence Data; D016366:Open Reading Frames; D016415:Sequence Alignment; D017386:Sequence Homology, Amino Acid
D018076:DNA, Complementary; D006148:Guanine Deaminase