Influence of OGDHC and SAMs on Epothilone Biosynthesis
|Keywords||Sorangium cellulosum epothilone biosynthesis α-Oxoglutaratedehydrogenase S-adenosylmethionine synthetase|
Epothilones, which are macrocyclic lactone anticancer compounds naturallyproduced by the myxobacterium Sorangium cellulosum. As the same as the anticancermechanisms of paclitaxel, epothilones can inhibit tumor cell growth effectively.Epothilones have simple molecular structure, better solubility in water, less side effects,wide antitumor spectrum compared with paclitaxel. At the same time, epothilones canbe produced in fermentation, it is a kind of new antitumor compounds have broaddevelopment potential. Athrough the main research of epothilone have changed fromimproving fermentation production to clinical experiments.In this paper, several inhibitors including NaClO and H2O2were used to study theinhibitory effects on the growth, enzyme activity and yield of epothilones inSorangium cellulosum. Activities of α-Oxoglutarate dehydrogenase in S. cellulosumwith different inhibitors were lower than the control in various degrees. Titers ofepothilones A and B were reduced90%and96%by0.4mM NaClO, while5mMH2O2reduced them by46%and68%. The result indicated that methylmalonyl-CoA asone of the precursors of epothilones mainly transformed by succinyl-CoAtransformation and α-Oxoglutarate dehydrogenase complex played an important rolein epothilones biosynthesis. In the study of the relationship betweenS-adenosylmethionine synthetase (SAMs) activity and the biosynthesis of epothilone.SAMs showed high activities in the period of the biosynthesis of epothilone. SAMsactivities and the yields of epothilones both reduced with the0.2mmol/LIndole-3-acetic acid addition and increased with the4mmol/L Sodiump-toluenesulfonate addition. Result showed that the SAMs activities were positivecorrelated with the yields of epothilone when the inhibitor or accelerator was added.SAMs play an important role in the biosynthesis of epothilone in Sorangiumcellulosums.When analysing the amount of gene expression, the expression level ofthioesterase was used to express epothilone synthesis, ogdcE1was used to show theexpression level of OGDHC.16S, as reference gene,was used to analyze expression ofOGDHC and SAMs in the process of epothilone synthesis. Experiments resultsshowed that the amount of epothilone expression reached the highest whenfermentating3and4days and decreased when existing OGDHC inhibitors, which was consistent with the amount of epothilone. The expression of ogdc reducedsubstantially when adding H2O2but unobviously when adding NaClO. The reasoncould be the different working points, NaClO works in the enzyme level, but H2O2works in transcriptional level. IAA brought to the reducing of expression amount ofSAMs and TE. Simultaneously, p-TSA-Na brought to the rise.It may because that IAArestrains the transcription of SAMs but p-TSA-Na promotes it.