(Below N is a link to NCBI taxonomic web page and E link to ESTHER at designed phylum.) > cellular organisms: NE > Eukaryota: NE > Viridiplantae: NE > Streptophyta: NE > Streptophytina: NE > Embryophyta: NE > Tracheophyta: NE > Euphyllophyta: NE > Spermatophyta: NE > Magnoliophyta: NE > Mesangiospermae: NE > eudicotyledons: NE > Gunneridae: NE > Pentapetalae: NE > rosids: NE > fabids: NE > Fabales: NE > Fabaceae: NE > Papilionoideae: NE > Phaseoleae: NE > Phaseolus: NE > Phaseolus vulgaris: NE
LegendThis sequence has been compared to family alignement (MSA) red => minority aminoacid blue => majority aminoacid color intensity => conservation rate title => sequence position(MSA position)aminoacid rate Catalytic site Catalytic site in the MSA MEYIVHRTVEVNGIKMHVAEKGEGPAVLFLHGFPELWYTWRHQILDLSSR GYHAVAPDLRGYGDTEAPASMSSYSCFDIVGDLVALIDLLGVDQVFLVAH DWGAIIGWYLCMFRPDRVKAYVCLSVPFWPRNPKVKPVDAMRALYGDDYY ICRFQEAGKAEGELAKNSTEEVLKKLLTNRTPGPPILQKEGMGSNLNTSM PLPSWLSLQDLKYYASKFEKTGFTGGLNYYRNINLNWELTAPWTGAQVKV PVKFITGDLDSVYTSLGMKNYIESGAFKKDVPNLEEVIVQEGVAHFNNQE AAEDVSNHIYDFIKNF
Reference
Title: Greatly enhancing the enantioselectivity of PvEH2, a Phaseolus vulgaris epoxide hydrolase, towards racemic 1,2-epoxyhexane via replacing its partial cap-loop Li C, Hu BC, Wen Z, Hu D, Liu YY, Chu Q, Wu MC Ref: Int J Biol Macromol, 156:225, 2020 : PubMed
To achieve the kinetic resolution and enantioconvergent hydrolysis of rac-1,2-epoxyhexane, the E value of PvEH2 was enhanced by substituting its partial cap-loop. Based on the experimental results reported previously and computer-aided analysis, the flexible and variable cap-loop, especially its middle segment, was speculated to be related to the catalytic properties of PvEH2. In view of this, four PvEH2's hybrids, Pv2St, Pv2Pv1, Pv2Vr1 and Pv2Vr2, were designed by substituting the middle segment ((190)EGMGSNLNTSMP(201)) of a cap-loop in PvEH2 with the corresponding ones in StEH, PvEH1, VrEH1 and VrEH2, respectively. Then, the hybrid-encoding genes, pv2st, pv2pv1, pv2vr1 and pv2vr2, were constructed by fusion PCR, and expressed in E. coli Rosetta(DE3). The expressed hybrid, Pv2St, displayed the highest specific activity of 35.3 U/mg protein towards rac-1,2-epoxyhexane. The corresponding transformant, E. coli/pv2st, exhibited the largest E value of 24.2, which was 11.5-fold that of E. coli/pveh2 expressing PvEH2. The scale-up kinetic resolution of 280 mM rac-1,2-epoxyhexane was carried out using 40 mg dry cells/mL of E. coli/pv2st at 25 degrees C for 4.5 h, retaining (S)-1,2-epoxyhexane with >99.5% ees and 36.9% yield. Additionally, the chemo-enzymatic enantioconvergent hydrolysis of rac-1,2-epoxyhexane using E. coli/pv2st followed by sulfuric acid produced (R)-hexane-1,2-diol with 73.0% eep and 86.5% yield.
