Chemical characterisation of disruptants of the Streptomyces coelicolor A3(2) actVI genes involved in actinorhodin biosynthesis

The actVI genetic region of Streptomyces coelicolor A3(2) is part of the bi osynthetic gene cluster of actinorhodin (ACT), the act cluster, consisting of six ORFs: ORFB, ORFA, ORF1, ORF2, ORF3, ORF4. A newly devised method of ACT detection with a combination of HPLC and LC/MS was applied to the analy sis of the disruptants of each ORE ACT was produced by those of ORFB, ORFA, ORF3, and ORF4. Instead of ACT, the ORF1 disruptant produced 3,8dihydroxy- 1-methylanthraquinone-2-carboxylic acid (DMAC) and aloesaponarin II as shun t products. The ORF2 disruptant gave 4-dihydro-9-hydrsxy-1-methyl-10-oxo-3- H-naphtho-[2,3-c]-pyran-3-(S)-acetic acid, (S)-DNPA. These results support our previous proposal for stereospecific pyran ring formation in the biosyn thesis of ACT, most importantly suggesting that the actVI-ORF2 product woul d recognize (S)-DNPA as a substrate for stereospecific reduction at C-15. T he disruptant of ORFA produced (S)-DNPA together with ACT, suggesting that actVI-ORFA might play a role such as stabilising the multicomponent, type I I PKS complex.