甲基转移酶
化学
链霉菌
生物化学
生物
遗传学
甲基化
DNA
细菌
作者
F Fawaz,George H. Jones
标识
DOI:10.1016/s0021-9258(18)68824-6
摘要
A methyltransferase, which utilizes 3-hydroxyanthranilic acid (HAA) as a substrate, has been purified to near homogeneity from 30-36-h mycelium of the bacterium Streptomyces antibioticus.The enzyme was obtained in approximately 20% yield with a purification of 130-fold.Polyacrylamide gel electrophoresis under denaturing conditions indicates that the enzyme is composed of a single subunit with M , of about 36,000.On chromatography in 0.5 M NaCl, the enzyme displays a molecular weight of about 37,000.The specific activity of the enzyme in s. antibioticue mycelium is maximal between 30 and 36 h following inoculation of galactose/glutamic acid medium and, at those times post-inoculation, the specific activity is essentially the same in extracts of mycelium obtained from cultures grown on glucose rather than galactose as the carbon source.The enzyme activity is stimulated by NazEDTA (in crude extracts) and by 2-mercaptoethanol and the methyltransferase shows a strong preference for HAA as substrate as compared with a number of HAA analogs.Thin layer chromatography of ethyl acetate extracts of large-scale incubation mixtures confirms that the product of the reaction is 4-methyl-3-hydroxyanthranilic acid.The reaction product was also a substrate for phenoxazinone synthase and was incorporated into actinomycin by S. antibioticus mycelium.Kinetic parameters for the methyltransferase reaction was determined.Considerable progress has been made in recent years in characterizing the enzymes which are involved in the biosynthesis of the antibiotic, actinomycin.Thus, Keller and coworkers (1) have described the isolation of an enzyme that activates 4-methyl-3-hydroxyanthranilic acid (MHA),' which serves as the initial substrate in the polymerization of the peptide chains of the antibiotic.More recently, Keller has reported the characterization of two actinomycin synthetases which modify and polymerize the amino acids in the actinomycin pentapeptide chains (2).There is strong evidence to suggest that it is the MHA-pentapeptides which are the substrates in the ultimate or penultimate step in the biosynthetic pathway, the synthesis of the phenoxazinone ring which comprises the actinomycin chromophore (3, 4).Phenoxazi-
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