Impact And Regulation Mechanism Of Quorum Sensing On Biosynthesis Of Lovastatin And Red Pigments In Monascus

BackgroundRecent some studies indicated that a phenomenon called quorum sensing has great potential value for enhancing the yield of secondary metabolite production in microorganism. In this study, we investigated linoleic acid-derived signaling molecules has an impact on biosynthesis of lovastatin and red pigments and mechanism of regulate process in Monascus.MethodsThe yield of lovastatin and red pigments were evaluated by high-performance liquid chromatography and ultraviolet spectrophotometry, respectively. Meanwhile, the dose and time-effects relationship of biosynthesis of lovastatin and red pigments treated with linoleic acid were discussed. The m RNA expression of regulatory gene mk H and lovastatin synthetic gene mk A were analyzed by Real Time PCR assay.Cyclic adenosine monophosphate(c AMP) and intracellular calcium ion in G protein signaling pathways were evaluatedenzyme linked immunosorbent assay and colorimetric method which the dose and time- content relationship were discussed. In addition, protein kinase A(PKA) and protein kinase C(PKC) activity were evaluated by non-radioactive protein kinase detection and phospholipase C activity were evaluated by enzyme-linked immunoassay, which of the dose and time- activity relationship were discussed.ResultsLinoleic acid(LA)has an effect that the production of lovastatin and red pigments were enhanced in a dose-dependent manner in Monascus. The low concentration of linoleic acid was advantageous to the production of red pigments and red pigments production was 5.6U/m L and increased 27% compared to the control culture at day 13 of the fermentation when LA was added to the production medium at concentrations of 64μM. In addition, a high concentration of linoleic acid was advantageous to the production of lovastatin and the production was 13.2mg/g and increased 26% compared to the control culture at day 13 of the fermentation when LA was added to the production medium at concentrations of 512μM. Real-time quantitative PCR results show that the high concentration of LA could up-regulate the expression of gene mk A and the mk A was transcribed at lower levels in cultures when low concentration of linoleic acid was added. The highest levels of gene mk A was transcribed at 512μM of LA added at 9 days and the lowest levels was transcribed was at 64μM of LA added at 7 days corresponding to an levels of 5.36- and 0.27-fold compared to the control, respectively. the mk H gene in linoleic acid- added cultures was expressed at higher levels compared to the control cultures at the premetaphase stage of the fermentations. The greatest increase obtained at day 9 when the transcriptional levels of mk H gene in the linoleic acid-supplemented cultures was 11.8-fold for 64μM and 7.41-fold 512μM compared with control cultures.The effects of linoleic acid on c AMP-PKA signal pathway in Monascus cultures lead to increase c AMP intracellular concentration in a dose-dependent and maintained at a high level. Meanwhile, protein kinase A(PKA) activity was significant increased in a LA dose-dependent and maintained at a high level which would be consistent with the change of the c AMP. Protein kinase C(PKC) activity was increased but intracellular calcium ion concentration and phospholipase C activity were no significant change compared with control cultures in Phosphatidylinositol signal pathway.ConclusionThe process of producing red pigment was changed upon exogenous addition of the linoleic acid to low cell density cultures of Monascus. The yield of red pigment inversely proportional but lovastatin directly proportional to enhance with the addition concentration of the linoleic acid and production of both were competitive within a range. Lovastatin synthetic gene mk A transcript levels was proportional to production of lovastatin but was inversely proportional to production of red pigment. The regulatory gene mk H transcript levels was related to yield of both of red pigment and lovastatin and the more active of secondary metabolic activity,the higher gene mk H transcript levels in Monascus. The effects of linoleic acid on c AMP-PKA signal pathway for yield of both of red pigment and lovastatin was significant in Monascus and phosphatidyl inositol-PKC signaling pathway has a influence.Howere, it was no significant effect on secondary metabolic activity that phosphatidyl inositol- Ca2+ signal pathway.