4 CONCLUSION
Mycobacterium could degrade the sidechain of PS to produce the
steroidal intermediates 9-OH-AD. In this study, the production of
9-OH-AD from PS biotransformation was significantly improved by
modulating the key gene expression of kshA1 and hsd4A in
mycobacterial mutants. The augment of gene hsd4A could
effectively promote the side-chain degradation of PS and decrease the
incomplete degradation products, DHBC, but increase the complete
degradation product of 4-androstene-3,17-dione (AD) due to lack of
3-Ketosteroid 9α-Hydroxylase KshA1 activities. Therefore, co-expression
of kshA1 and hsd4A in an appropriate pattern in
mycobacterial could optimize their expression level and achieve a better
balance of the Hsd4A and KshA1 activity. Further decreasing the PS
conversion rate could be best for the action of the Hsd4A and KshA1
activities to transfer the metabolic flux to the target product of
9-OH-AD and decrease the byproducts formation. Finally, the yield and
purity of 9-OH-AD in PS transformation by HK86 K-A mutant were increased
from 22.18 g/L and 77.13% to 28.27 g/L and 87.84%, respectively, with
a molar yield of 78.32%.