title: Purification and characterization of GTP cyclohydrolase I from Streptomyces tubercidicus, a producer of tubercidin.
abstract: GTP cyclohydrolase I catalyzing the first reaction in the biosynthesis of pterin moiety of folic acid in bacteria, was purified from Streptomyces tubercidicus by at least 203-fold with a yield of 32% to apparent homogeneity, using ammonium sulfate fractionation, DEAE-cellulose, Sepharose CL-6B, and hydroxylapatite column chromatography. The molecular weight of the native enzyme was estimated to be 230,000 daltons by gel permeation chromatography. The purified enzyme gave a single band on sodium dodesyl sulfate-polyacrylamide gel electrophoresis and its molecular weight was apparently 58,000 daltons. These results indicate that the enzyme consists of four subunits with the same molecular weight. The K(m) and Vmax values for GTP of the purified enzyme were determined to be 80 microM and 90 nmol/min (mg protein), respectively. The optimum pH and temperature for the enzyme reaction were pH 7.5-8.5 and 40-42 degrees C, respectively. Coenzyme or metal ion was not required for the enzyme activity. The enzyme activity was inhibited by most divalent cations, while it was slightly activated by potassium ion. In case of nucleotides, CTP, GMP, GDP, and UTP inhibited enzyme activity, among which GDP exhibited the strongest inhibitory effect.
authors.affiliation: Department of Pharmacy, Chosun University, Kwangju, Korea.
authors.forename: J C
authors.initials: JC
authors.lastname: Yoo
authors.forename: J M
authors.initials: JM
authors.lastname: Han
authors.forename: O H
authors.initials: OH
authors.lastname: Ko
authors.forename: H J
authors.initials: HJ
authors.lastname: Bang
mesh_terms: D000900:Anti-Bacterial Agents / Q000737:chemistry / Q000302:isolation & purification; D006136:GTP Cyclohydrolase / Q000737:chemistry* / Q000302:isolation & purification* / Q000493:pharmacokinetics; D008970:Molecular Weight; D011741:Pyrimidine Nucleosides / Q000737:chemistry / Q000494:pharmacology; D013302:Streptomyces / Q000737:chemistry*; D014372:Tubercidin / Q000096:biosynthesis
chemical_list: D000900:Anti-Bacterial Agents; D011741:Pyrimidine Nucleosides; D006136:GTP Cyclohydrolase; D014372:Tubercidin