title: Characterization of representative rpoB gene mutations leading to a significant change in toyocamycin production of Streptomyces diastatochromogenes 1628.
abstract: Modification of enzymes involved in transcription- or translation-processes is an interesting way to increase secondary metabolite production in Streptomycetes. However, application of such methods has not been widely described for strains which produce nucleoside antibiotics. The nucleoside antibiotic toyocamycin (TM) is produced by Streptomyces diastatochromogenes 1628. For improving TM production in S. diastatochromogenes 1628, the strain was spread on rifamycin-resistant (Rif(r)) medium. Several spontaneous mutants were obtained with mutations in the rpoB gene which encodes a RNA polymerase β-subunit. The mutants which showed increased TM production were detected at a frequency of 7.5 % among the total Rif(r) mutants. Mutant 1628-T15 harboring amino acid substitution His437Arg was the best TM producer with a 4.5-fold increase in comparison to that of the wild-type strain. The worst producer was mutant 1628-T62 which also showed a poor sporulation behavior. RT-PCR was performed to study the transcription levels of the TM biosynthetic gene toyG in the parental strain as well as in mutants 1628-T15 and 1628-T62. The transcriptional level of toyG was higher in mutant 1628-T15 than that in parental strain 1628, while much lower in mutant 1628-T62. In mutant strain 1628-T62 the expression of adpA sd gene, which is required for morphological differentiation, was also much lower. Our studies also indicate that the introduction of mutations into rpoB is an effective strategy to improve the production of TM which is an important nucleoside antibiotic.
authors.affiliation: Zhejiang Provincial Key Laboratory of Biometrology and Inspection and Quarantine, College of Life Sciences, China Jiliang University, 310018, Hangzhou, Zhejiang Province, China.
authors.forename: Zheng
authors.initials: Z
authors.lastname: Ma
authors.affiliation: Zhejiang Provincial Key Laboratory of Biometrology and Inspection and Quarantine, College of Life Sciences, China Jiliang University, 310018, Hangzhou, Zhejiang Province, China.
authors.forename: Shuai
authors.initials: S
authors.lastname: Luo
authors.affiliation: Zhejiang Provincial Key Laboratory of Biometrology and Inspection and Quarantine, College of Life Sciences, China Jiliang University, 310018, Hangzhou, Zhejiang Province, China.
authors.forename: Xianhao
authors.initials: X
authors.lastname: Xu
authors.affiliation: Pharmazeutische Biologie und Biotechnologie, Institut für Pharmazeutische Wissenschaften, Albert-Ludwigs-Universität Freiburg im Breisgau, Stefan-Meier-Straße, 19, 79104, Freiburg, Germany.
authors.forename: Andreas
authors.initials: A
authors.lastname: Bechthold
authors.affiliation: Zhejiang Provincial Key Laboratory of Biometrology and Inspection and Quarantine, College of Life Sciences, China Jiliang University, 310018, Hangzhou, Zhejiang Province, China. yuxiaoping19630306@163.com.
authors.forename: Xiaoping
authors.initials: X
authors.lastname: Yu
mesh_terms: D000903:Antibiotics, Antineoplastic / Q000096:biosynthesis*; D001426:Bacterial Proteins / Q000235:genetics*; D053898:Biosynthetic Pathways / Q000235:genetics; D012321:DNA-Directed RNA Polymerases / Q000235:genetics*; D009154:Mutation / Q000235:genetics*; D012294:Rifamycins / Q000494:pharmacology; D013171:Spores, Bacterial / Q000235:genetics; D013302:Streptomyces / Q000187:drug effects / Q000235:genetics* / Q000378:metabolism*; D014127:Toyocamycin / Q000096:biosynthesis*
chemical_list: D000903:Antibiotics, Antineoplastic; D001426:Bacterial Proteins; D012294:Rifamycins; D012321:DNA-Directed RNA Polymerases; C105078:RNA polymerase beta subunit; D014127:Toyocamycin
keywords: Diastatochromogenes; GUS; Rifamycin-resistant mutant; RpoB gene; Streptomyces; Toyocamycin