Fishman S E, Cox K, Larson J L, Reynolds P A, Seno E T, Yeh W K, Van Frank R, Hershberger C L
Lilly Corporate Center, Indianapolis, IN 46285.
Proc Natl Acad Sci U S A. 1987 Dec;84(23):8248-52. doi: 10.1073/pnas.84.23.8248.
Macrocin-O-methyltransferase (MacOMeTase) catalyzes the final enzymatic step in the biosynthesis of tylosin in Streptomyces fradiae. A 44-base mixed oligonucleotide probe containing only guanosine and cytidine in the third position of degenerate codons was synthesized based on the amino acid sequence of the amino terminus of MacOMeTase. Plaque blot hybridization to a bacteriophage lambda library and colony blot hybridization to a cosmid library of S. fradiae DNA identified recombinants that contained overlapping fragments of chromosomal DNA. The nucleotide sequence of the cloned DNA verified that the DNA contained the coding sequence for MacOMeTase. Recombinant plasmids transformed mutants blocked in tylosin biosynthesis and complemented tylF (the structural gene for MacOMeTase) and tyl mutations of eight other classes.
大环菌素 - O - 甲基转移酶（MacOMeTase）催化弗氏链霉菌中泰乐菌素生物合成的最后一步酶促反应。基于MacOMeTase氨基末端的氨基酸序列，合成了一个44个碱基的混合寡核苷酸探针，该探针在简并密码子的第三位仅含有鸟苷和胞苷。与弗氏链霉菌DNA的λ噬菌体文库进行噬菌斑杂交以及与粘粒文库进行菌落杂交，鉴定出含有染色体DNA重叠片段的重组体。克隆DNA的核苷酸序列证实该DNA包含MacOMeTase的编码序列。重组质粒转化在泰乐菌素生物合成中受阻的突变体，并互补了tylF（MacOMeTase的结构基因）和其他八类tyl突变。
