Biological Effects And Regulation Of MBD In Comamonas Testosteroni

Meso-butanediol dehydrogenase(MBD)is an enzyme in Comamonas testosteroni(C.T.)ATCC11996.As it can catalyze the reaction to produce a large amount of bioproducts,acetoin(AC)and meso-2,3-butanediol(meso-2,3-BD),has become a focus of experts and scholars.In this paper,MBD was used as the research object to explore the biological effects and gene regulation of steroid hormones in the testicular bacterium of C.testosteroni.Through hormone induction,total RNA extraction,whole transcriptome sequencing,sequencing information alignment,gene amplification,construction of gene expression vector,exogenous protein expression,protein affinity chromatography purification,gene knockout,plasmid co-transformation,high performance liquid chromatography,ELISA,and other experimental methods,the results show that:A total of 4,835 genes in C.T.strains induced by testosterone were different at the transcriptional level,and there were 110 significant differential genes.The Log2 value of MBD gene difference reached 1.80,which was 3.48 times different from that before induction.By constructing MBD knockout C.T.strain and using high performance liquid chromatography to detect its degradation ability to testosterone,methyltestosterone,estrone,estradiol,and other steroid hormones,studies have shown that after knockout of the meso-butanediol dehydrogenase gene in C.T.the degradation rate of testosterone is 27.80%,which is 47.40% lower than that of wild type;the degradation rate of methyltestosterone in the knockout strain was 24.10%,which was 18.20% lower than that of the wild type;the degradation rate of estrone by the knockout strain was 14.80%,which was lower than that of the wild type.16.20%;the degradation rate of estradiol by the knockout strain was 19.30%,which was 5.60% lower than that of the wild type.The MBD co-transformation ELISA assay showed that the expression of MBD was decreased in LuxR and MBD plasmid co-transformants,the expression of MBD in LysR and MBD plasmid co-transformants was increased,and the expression of MBD in TetR and MBD plasmid co-transformants was not significantly different.The experimental results in this study indicate that MBD plays an important biological role in the metabolism of testosterone and other steroid hormones in C.testosteroni.Two transcriptional regulators LuxR and LysR in C.testosteroni have a regulatory effect on MBD,of which LuxR It has a down-regulation effect,and LysR has an up-regulation effect.The results provide an important theoretical and experimental basis for the study of steroid hormone metabolism pathways and gene regulation in C.T..