Purification and characterization of a novel nucleoside phosphorylase from a Klebsiella sp. and its use in the enzymatic production of adenine arabinoside.
An adenosine-assimilating bacterium, Klebsiella sp. strain LF1202, inducibly formed a novel nucleoside phosphorylase which acted on both purine and pyrimidine nucleosides when the cells were cultured in medium containing adenosine as a sole source of carbon and nitrogen. The enzyme was purified (approximately 83-fold, with a 17% activity yield) to the homogeneous state by polyacrylamide gel electrophoresis. The molecular weight of the purified enzyme was calculated to be 125,000 by gel filtration of Sephadex G-200 column chromatography, although the enzyme migrated as a single protein band with a molecular weight of 25,000 on sodium dodecyl sulfate (SDS)-polyacrylamide gel electrophoresis; thus, it was thought to consist of five identical subunits. Besides purine nucleosides (adenosine, inosine, and guanosine), the purified enzyme also acted on pyrimidine nucleosides such as uridine, 2'-deoxyuridine, and thymidine. The purified enzyme catalyzed the synthesis of adenine arabinoside, a selective antiviral pharmaceutic agent, from uridine arabinoside and adenine.
Ling
F
F
Research Institute for Food Science, Kyoto University, Japan.
Inoue
Y
Y
Kimura
A
A
D000225:Adenine / Q000378:metabolism; D002848:Chromatography, DEAE-Cellulose; D002850:Chromatography, Gel; D004591:Electrophoresis, Polyacrylamide Gel; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D007709:Klebsiella / Q000201:enzymology*; D008970:Molecular Weight; D010430:Pentosyltransferases / Q000737:chemistry / Q000302:isolation & purification* / Q000378:metabolism; D011684:Purine Nucleosides / Q000378:metabolism; D011741:Pyrimidine Nucleosides / Q000378:metabolism; D013379:Substrate Specificity; D013696:Temperature; D014740:Vidarabine / Q000096:biosynthesis*
D011684:Purine Nucleosides; D011741:Pyrimidine Nucleosides; D010430:Pentosyltransferases; C067995:nucleoside phosphorylase; D014740:Vidarabine; D000225:Adenine