A novel GDP-mannose mannosyl hydrolase shares homology with the MutT family of enzymes.
The product of the Escherichia coli orf1.9, or yefc, gene (GenBank accession number L11721) has been expressed under the control of a T7 promoter, purified to apparent homogeneity, and identified as a novel enzyme that hydrolyzes GDP-mannose or GDP-glucose to GDP and the respective hexose. The enzyme has little or no activity on other nucleotides, dinucleotides, nucleotide sugars, or sugar phosphates. It has a pH optimum between 9.0 and 9.5, a Km of 0.3 mM, and a Vmax of 1.6 mumol min-1 mg-1 for GDP-mannose, and it requires divalent cations for activity. This enzyme of 160 amino acids (M(r) = 18, 405) contains the consensus sequence GX(I/L/V)(E/Q)(X)2ET(X)6R(X)4E(X)2(I/L), characteristic of the MutT family of proteins and previously shown to form part of the nucleotide-binding site of MutT (Frick, D. N., Weber, D. J., Abeygunawardana, C., Gittis, A. G., Bessman, M. J., and Mildvan, A. S. (1995) Biochemistry 34, 5577-5586). A comparison of the enzymatic reactions catalyzed by the GDP-mannose mannosyl hydrolase and the other enzymes of the MutT family suggests that the consensus signature sequence designates a novel nucleoside diphosphate binding site and catalytic motif.
Frick
D N
DN
Department of Biology, Johns Hopkins University, Baltimore, Maryland 21218, USA.
Townsend
B D
BD
Bessman
M J
MJ
D000595:Amino Acid Sequence; D001419:Bacteria / Q000737:chemistry / Q000235:genetics; D001426:Bacterial Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D001483:Base Sequence; D002876:Chromosomes, Bacterial; D003001:Cloning, Molecular; D017931:DNA Primers; D004926:Escherichia coli / Q000201:enzymology* / Q000235:genetics; D029968:Escherichia coli Proteins; D006026:Glycoside Hydrolases / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D006801:Humans; D007700:Kinetics; D008969:Molecular Sequence Data; D010744:Phosphoric Monoester Hydrolases / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D011755:Pyrophosphatases; D011994:Recombinant Proteins / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism; D017386:Sequence Homology, Amino Acid; D013379:Substrate Specificity
D001426:Bacterial Proteins; D017931:DNA Primers; D029968:Escherichia coli Proteins; D011994:Recombinant Proteins; D010744:Phosphoric Monoester Hydrolases; D006026:Glycoside Hydrolases; C096402:yefc protein, E coli; D011755:Pyrophosphatases; C052211:mutT protein, E coli