In vitro catalytic production of prostaglandin F2α using arachidonic acid
As the important lipid mediators,prostaglandin F2α(PGF2α)performs a wide range of physiological activities.To achieve the green biosynthesis of PGF2α,pET30a-TbPGFS,pET30a-MmPGHS and pET30a-GvPGHS vectors were constructed and expressed in Escherichia coli BL21(DE3),respectively,and the induction expression conditions of isopropyl-β-D-thiogalactopyranoside(IPTG)were optimized to improve the expression of the recombinant protein in E.coli.In addition,the recombinant protein obtained under the optimal conditions were used as a catalyst to catalyze the synthesis of PGF2α from arachidonic acid.The results showed that MmPGHS protein was not expressed in E.coli.However,GvPGHS protein performed the optimized expression under the conditions of IPTG concentration 0.2 mmol/L,induction temperature 30 ℃ and induction time 2 h,the TbPGFS protein performed the optimized expression under the conditions of IPTG concentration 0.2 mmol/L,induction temperature 30 ℃ and induction time 6 h.Under the above optimized conditions,PGF2α was not detected by the two-enzyme coupled catalysis system composed of crude extract of GvPGHS and TbPGFS.However,arachidonic acid was converted into PGH2 by GvPGHS,which was further reduced to PGF2α by SnCl2 under the enzyme-coupled chemical catalysis.In conclusion,In vitro enzymatic reaction combined with chemical method can efficiently convert arachidonic acid to produce PGF2α.
arachidonic acidprostaglandin F2αin vitro catalysisprostaglandin H synthase
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