Five gene products are required for assembly of the central pyrrole moiety of coumermycin A(1)
| Autoři | |
|---|---|
| Rok publikování | 2013 |
| Druh | Článek v odborném periodiku |
| Časopis / Zdroj | JOURNAL OF INDUSTRIAL MICROBIOLOGY & BIOTECHNOLOGY |
| Fakulta / Pracoviště MU | |
| Citace | |
| www | Full Text |
| Doi | http://dx.doi.org/10.1007/s10295-013-1266-6 |
| Obor | Biotechnologie a bionika |
| Klíčová slova | Streptomyces; Coumermycin; Pyrrole; Biosynthesis; Heterologous expression |
| Popis | Coumermycin A(1) is an aminocoumarin antibiotic produced by Streptomyces rishiriensis. It exhibits potent antibacterial and anticancer activity. The coumermycin A(1) molecule contains two terminal 5-methyl-pyrrole-2-carboxylic acid moieties and one central 3-methylpyrrole-2,4-dicarboxylic acid moiety (CPM). While the biosynthesis of the terminal moieties has been elucidated in detail, the pathway leading to the CPM remains poorly understood. In this work, the minimal set of genes required for the generation of the CPM scaffold was identified. It comprises the five genes couR1, couR2a, couR2b, couR3, and couR4 which are grouped together in a contiguous 4.7 kb region within the coumermycin A(1) biosynthetic gene cluster. The DNA fragment containing these genes was cloned into an expression plasmid and heterologously expressed in Streptomyces coelicolor M1146. Thereupon, the formation of CPM could be shown by HPLC and by HPLC-MS/MS, in comparison to an authentic CPM standard. This proves that the genes couR1-couR4 are sufficient to direct the biosynthesis of CPM, and that the adjacent genes couR5 and couR6 are not required for this pathway. The enzyme CouR3 was expressed in Escherichia coli and purified to near homogeneity. The protein exhibited an ATPase activity similar to that reported for its close ortholog, the threonine kinase PduX. However, we could not show a threonine kinase activity of CouR3, and; therefore, the substrate of CouR3 in CPM biosynthesis is still unknown and may be different from threonine. |
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