Preparation of lipase with improved ester synthesis activity by using surfactants

Abstract

The present disclosure discloses a method for preparing lipase with high ester synthesis activity by using surfactant, belonging to the field of enzyme engineering. The present disclosure provides a method for obtaining a lipase with high ester synthesis activity by adding different surfactants with different concentrations in a lipase aqueous solution and then freeze-drying. The lipase meets the requirement in non-aqueous catalysis. Mixing a variety of lipase with no ester synthesis activity or low activity at suitable concentration and an appropriate concentration of the surfactant in the solution can produces lipases with significantly improved ester synthesis activity, meanwhile changing the hydrolytic activity of the lipase. Increased ester synthesis activity makes lipase more suitable for industrial applications.

What is claimed is: 
     
         1 . A method for increasing ester synthesis activity of lipase, comprising adding surfactants directly to aqueous solution of soluble lipase and mixing afterwards. 
     
     
         2 . The method for increasing the ester synthesis activity of lipase according to  claim 1 , wherein the aqueous solution of the lipase is obtained by dissolving a lipase powder directly in the aqueous solution. 
     
     
         3 . The method for increasing the ester synthesis activity of lipase according to  claim 1 , wherein the aqueous solution is phosphate buffer saline or Tris-HCl buffer. 
     
     
         4 . The method for increasing the ester synthesis activity of lipase according to  claim 1 , wherein the surfactant is one or a combination of zwitterionic surfactants, nonionic surfactants and cationic surfactants. 
     
     
         5 . The method for increasing the ester synthesis activity of lipase according to  claim 1 , wherein the soluble lipase comprises  Rhizopus chinesis  lipase r27RCL,  Rhizopus oryzae  lipase ROL,  Pseudomonas cepacia  lipase PCL, or  Candida antarctica  lipase CALB. 
     
     
         6 . The method for increasing the ester synthesis activity of lipase according to  claim 1 , wherein the soluble lipase is a lipase that is heterologously expressed, a lipase expressed by a recombinant microorganism, or any other lipases that have lower or no ester synthesis activity relative to a wild-type lipase. 
     
     
         7 . The method for increasing the ester synthesis activity of lipase according to  claim 1 , wherein protein concentration of pure lipase in the aqueous solution of the soluble lipase is between 0.1 and 0.4 mg·mL −1 . 
     
     
         8 . The method for increasing the ester synthesis activity of lipase according to  claim 4 , wherein the zwitterionic surfactants comprises 3-[3-(cholamidopropyl) dimethylamino] propanesulfonic acid internal salt, polyethylene glycol octyl Phenyl ether, 1-myristyl 2-acyl-cis-propyltriyl-phosphatidylcholine, 1,2-dihexanoyl lecithin, 1-myristyl-2-acyl-cis-propyltri-2-acyl-cis-propyltriate-phosphatidylcholine, 1-myristyl-2-acyl-cis-propyltriyl-phosphonic acid-1-glycerol, 1-myrist 2-acyl-cis-propyltri-phospho-1-glycerol or 1,2-diheptanoyl-S-glyceryl, Glycerol-3-phosphorylcholine; the nonionic surfactant is dodecyl-ß- D -maltoside, octylglycol monobutyl dodecyl ester, decyl-ß- D -maltose Glycoside, undecyl-ß- D -maltoside, octyl-ß- D -glucopyranoside or polyoxypropylene glycol monethylether. 
     
     
         9 . The method for increasing the ester synthesis activity of lipase according to  claim 4 , wherein the nonionic surfactants comprises n-dodecyl-ß- D -maltopyranoside, octaethylene glycol monododecyl ether, n-decyl-ß- D -maltopyranoside, n-undecyl-ß- D -maltopyranoside, n-octyl-ß- D -glucopyranoside or octaethylene glycol monodecyl ether. 
     
     
         10 . The method for increasing the ester synthesis activity of lipase according to  claim 1 , wherein final concentration of surfactant is ranging from 0.1 times to 200 times critical micelle concentration and above. 
     
     
         11 . The method for increasing the ester synthesis activity of lipase according to  claim 1 , wherein the addition of the surfactant is 1-500 mM or higher at final concentration. 
     
     
         12 . A lipase preparation with improved ester synthesis activity in non-aqueous phase, comprising a lipase preparation obtained by dissolving the lipase in an aqueous solution and then adding surfactant directly to the solution followed by removing water. 
     
     
         13 . A method comprising adding the lipase preparation according to  claim 12  as a catalyst in catalyzing ester synthesis reaction in non-aqueous phase; or adding the lipase preparation according to  claim 12  as a catalyst in catalyzing a reaction in field of foodstuffs, chemistry, biology, preparation of medicine, or environment or petroleum field.