Expression of Candida antarctica lipase B in Pichia pastoris and Studying of Enzyme Properties

To improve expression efficiency of recombinant Candida antarctica lipase B(CALB) in methylotrophic yeast Pichia pastoris GS115,the DNA sequence encoding CALB was designed and synthesized based on the codon bias of P.pastoris.The new CALB gene was cloned into pPIC9K,yielding pPIC9K-CALB,and transformed into P.pastoris GS115.In shake-flask cultivation,the enzyme activity was 46 U/mL after induction by methanol for 120 h.The CALB was purified by metal chelate chromatography by 5.23 folds.Deglycosylation showed the recombinant CALB was 5 ku bigger than wild type CALB.The optimal pH and temperature of this recombinant enzyme were pH 6.5 and 30°C,respectively.CALB was relatively stable at temperatures below 40 ℃ within a pH range from 5.0 to 8.0.Ca2+,Zn2+,Mn2+ conduced to enzyme activity,while Cu2+,Ag+,Fe3+ inhibited it.