Patent ID: 8227215

Claim:
A method for biocatalytic synthesis of a structured lipid comprising the following steps: (a) providing a polypeptide having a palmitase activity, said polypeptide being selected from the group consisting of isolated, synthetic and recombinant polypeptides having a palmitase activity and said polypeptide either i. being encoded by a nucleic acid comprising a nucleic acid sequence having at least 85% sequence identity to full length SEQ ID NO:1, and wherein the nucleic acid encodes at least one polypeptide having palmitase activity and the polypeptide having at least one amino acid residue change recited below in Table A, or ii. having at least 85% sequence identity to full length SEQ ID NO:2, and having at least one amino acid residue change recited in Table A, TABLE A Amino Original New Acid Amino Amino Residue Acid Acid 27 G Q 45 G L 54 S H 61 D A E 62 V A G 72 R E K S T 74 F I 83 V M 96 A I 98 G L 116 E A F H L Q Q R, H140R R S S T V 117 L R, I161L 120 K L 133 S A 147 I F 151 I A G H 160 P T 172 R Q 212 S R 213 K T 214 T V N R Y 215 G A H 216 A V C L or iii. having at least 85% sequence identity to full length of SEQ ID NO:2 and comprising at least one amino acid residue modification D61A; D61E; R72E; R72K; E116A; E116Q; E116R; E116T; E116V; S133A; I151G; I151A; V163R; or D164R (b) providing a composition comprising a triacylglyceride; (c) contacting the polypeptide of step (a) with the composition of step (b) under conditions wherein the polypeptide hydrolyzes a palmitic acid residue at the Sn2 position of the triacylglyceride, thereby producing a 1,3-diacylglyceride; (d) providing an R1 ester, wherein the R1 ester comprises a fatty acid of lower saturation than said hydrolyzed palmitic acid residue; (e) providing an R1-specific hydrolase, and (f) contacting the 1,3-diacylglyceride of step (c) with the R1 ester of step (d) and the R1-specific hydrolase of step (e) under conditions wherein the R1-specific hydrolase catalyzes esterification of the Sn2 position, thereby producing the structured lipid.