Gaisser S, Böhm G A, Doumith M, Raynal M C, Dhillon N, Cortés J, Leadlay P F
University of Cambridge, Department of Biochemistry, UK.
Mol Gen Genet. 1998 Apr;258(1-2):78-88. doi: 10.1007/s004380050709.
The gene cluster (ery) governing the biosynthesis of the macrolide antibiotic erythromycin A by Saccharopolyspora erythraea contains, in addition to the eryA genes encoding the polyketide synthase, two regions containing genes for later steps in the pathway. The region 5' of eryA that lies between the known genes ermE (encoding the erythromycin resistance methyltransferase) and eryBIII (encoding a putative S-adenosylmethionine-dependent methyltransferase), and that contains the gene eryBI (orf2), has now been sequenced. The inferred product of the eryBI gene shows striking sequence similarity to authentic beta-glucosidases. Specific mutants were created in eryBI, and the resulting strains were found to synthesise erythromycin A, showing that this gene, despite its position in the biosynthetic gene cluster, is not essential for erythromycin biosynthesis. A mutant in eryBIII and a double mutant in eryBI and eryBIII were obtained and the analysis of novel erythromycins produced by these strains confirmed the proposed function of EryBIII as a C-methyltransferase. Also, a chromosomal mutant was constructed for the previously sequenced ORF19 and shown to accumulate erythronolide B, as expected for an eryB mutant and consistent with its proposed role as an epimerase in dTDP-mycarose biosynthesis.
由糖多孢红霉菌合成大环内酯类抗生素红霉素A的基因簇（ery），除了包含编码聚酮合酶的eryA基因外，还含有两个区域，其中的基因参与该途径的后续步骤。eryA 5'端位于已知基因ermE（编码红霉素抗性甲基转移酶）和eryBIII（编码一种假定的依赖S-腺苷甲硫氨酸的甲基转移酶）之间，且包含基因eryBI（orf2），现已完成测序。eryBI基因的推断产物与真正的β-葡萄糖苷酶具有显著的序列相似性。在eryBI中构建了特定突变体，结果发现所得菌株能合成红霉素A，这表明该基因尽管位于生物合成基因簇中，但对红霉素生物合成并非必需。获得了eryBIII突变体以及eryBI和eryBIII双突变体，对这些菌株产生的新型红霉素的分析证实了EryBIII作为C-甲基转移酶的推测功能。此外，针对先前测序的ORF19构建了染色体突变体，结果显示其积累了红霉内酯B，这与eryB突变体预期一致，也与其在dTDP- mycose生物合成中作为差向异构酶的推测作用相符。
