Cloning, sequencing, and expression of the adenosylcobalamin-dependent ribonucleotide reductase from Lactobacillus leichmannii.
Ribonucleoside-triphosphate reductase (RTPR, EC 1.17.4.2) from Lactobacillus leichmannii, a monomeric adenosylcobalamin-requiring enzyme, catalyzes the conversion of nucleoside triphosphates to deoxynucleoside triphosphates. The gene for this enzyme has been cloned and sequenced. In contrast to expectations based on mechanistic considerations, there is no statistically significant sequence homology with the Escherichia coli reductase that requires a dinuclear-iron center and tyrosyl radical cofactor. The RTPR has been overexpressed and purified to homogeneity, yielding 90 mg of protein from 2.5 g of bacteria. Initial characterization of the recombinant RTPR indicates that its properties are identical to those of the RTPR isolated from L. leichmannii.
Booker
S
S
Department of Chemistry, Massachusetts Institute of Technology, Cambridge 02139.
Stubbe
J
J
D000595:Amino Acid Sequence; D001483:Base Sequence; D002845:Chromatography; D002848:Chromatography, DEAE-Cellulose; D003001:Cloning, Molecular; D003038:Cobamides / Q000494:pharmacology*; D017886:Durapatite; D004926:Escherichia coli / Q000235:genetics; D015870:Gene Expression; D006882:Hydroxyapatites; D007700:Kinetics; D007778:Lactobacillus / Q000201:enzymology* / Q000235:genetics; D008969:Molecular Sequence Data; D009838:Oligodeoxyribonucleotides; D010449:Peptide Mapping; D016133:Polymerase Chain Reaction; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000302:isolation & purification / Q000378:metabolism; D012264:Ribonucleotide Reductases / Q000096:biosynthesis* / Q000302:isolation & purification / Q000378:metabolism; D017386:Sequence Homology, Amino Acid
D003038:Cobamides; D006882:Hydroxyapatites; D009838:Oligodeoxyribonucleotides; D011994:Recombinant Proteins; D017886:Durapatite; D012264:Ribonucleotide Reductases; C000913:cobamamide