Method of Preparing Piceatannol Using Bacterial Cytochrome P450 and Composition Therefor

Abstract

Provided is a method of preparing piceatannol, and more particularly, to a method of preparing piceatannol from resveratrol using bacterial cytochrome P450 BM3 (CYP102A1) or mutants thereof, and a composition and a kit therefor.

1 . A composition for a catalyst in the reaction of preparing piceatannol from resveratrol, the composition comprising at least one selected from a group consisting of wild-type CYP102A1 and mutants of CYP102A1. 
     
     
         2 . The composition of  claim 1 , wherein the mutants of CYP102A1 are prepared by at least one selected from a group consisting of: substituting 47 th  amino acid arginine (R) of wild-type CYP102A1 with one amino acid selected from a group consisting of alanine, valine, leucine, isoleucine, proline, methionine, phenylalanine, and tryptophan, substituting 51 st  amino acid tyrosine(Y) of wild-type CYP102A1 with one amino acid selected from a group consisting of phenylalanine, alanine, valine, leucine, isoleucine, proline, methionine, tryptophan, substituting 64 th  amino acid glutamic acid (E) of wild-type CYP102A1 with one amino acid selected from a group consisting of glycine, serine, threonine, cysteine, tyrosine, asparagine, and glutamine, substituting 74 th  amino acid alanine (A) of wild-type CYP102A1 with one amino acid selected from a group consisting of glycine, serine, threonine, cysteine, tyrosine, asparagine, and glutamine, substituting 81 st  amino acid phenylalanine (F) of wild-type CYP102A1 with one amino acid selected from a group consisting of alanine, valine, leucine, isoleucine, proline, methionine, and tryptophan, substituting 86 th  amino acid leucine (L) of wild-type CYP102A1 with one amino acid selected from a group consisting of alanine, valine, isoleucine, proline, methionine, phenylalanine, and tryptophan, substituting 87 th  amino acid phenylalanine (F) of wild-type CYP102A1 with one amino acid selected from a group consisting of alanine, valine, leucine, isoleucine, proline, methionine, and tryptophan, substituting 143 rd  amino acid glutamic acid (E) of wild-type CYP102A1 with one amino acid selected from a group consisting of glycine, serine, threonine, cysteine, tyrosine, asparagine, and glutamine, substituting 188 th  amino acid leucine (L) of wild-type CYP102A1 with one amino acid selected from a group consisting of glycine, serine, threonine, cysteine, tyrosine, asparagine, and glutamine, and substituting 267 th  amino acid glutamic acid (E) of wild-type CYP102A1 with one amino acid selected from a group consisting of alanine, valine, leucine, isoleucine, proline, methionine, phenylalanine, and tryptophan. 
     
     
         3 . The composition of  claim 2 , wherein the mutants of CYP102A1 are prepared by at least one selected from a group consisting of: substituting 47 th  amino acid arginine (R) of wild-type CYP102A1 with leucine (L), substituting 51 st  amino acid tyrosine(Y) of wild-type CYP102A1 with phenylalanine, substituting 64 th  amino acid glutamic acid (E) of wild-type CYP102A1 with glycine (G), substituting 74 th  amino acid alanine (A) of wild-type CYP102A1 with glycine (G), substituting 81 st  amino acid phenylalanine (F) of wild-type CYP102A1 with isoleucine (I), substituting 86 th  amino acid leucine (L) of wild-type CYP102A1 with isoleucine (I), substituting 87 th  amino acid phenylalanine (F) of wild-type CYP102A1 with valine (V), substituting 143 rd  amino acid glutamic acid (E) of wild-type CYP102A1 with glycine (G), substituting 188 th  amino acid leucine (L) of wild-type CYP102A1 with glutamine (Q), and substituting 267 th  amino acid glutamic acid (E) of wild-type CYP102A1 with valine (V). 
     
     
         4 . The composition of  claim 1 , wherein the mutants of CYP102A1 comprises amino acid substitution sites of wild-type CYP102A1 selected from a group consisting of F87A, R47L/Y51 F, A74G/F87V/L188Q, R47L/L86I/L188Q, R47L/F87V/L188Q, R47L/F87V/L188Q/E267V, R47L/L86I/L188Q/E267V, R47L/L86I/F87V/L188Q, R47L/F87V/E143G/L188Q/E267V, R47L/E64G/F87V/E143G/L188Q/E267V, R47L/F81I/F87V/E143G/L188Q/E267V, and R47L/E64G/F81I/F87V/E143G/L188Q/E267V. 
     
     
         5 . The composition of  claim 1 , wherein the resveratrol is trans-resveratrol. 
     
     
         6 . A method of preparing piceatannol, the method comprising reacting at least one enzyme selected from a group consisting of wild-type CYP102A1 and mutants of CYP102A1 with resveratrol. 
     
     
         7 . The method of  claim 6 , further comprising adding an NADPH-generating system. 
     
     
         8 . The method of  claim 7 , wherein the NADPH-generating system comprises glucose 6-phosphate, NADP+, and yeast glucose 6-phosphate. 
     
     
         9 . The method of  claim 6 , wherein the resveratrol is trans-resveratrol. 
     
     
         10 . The method of  claim 6 , wherein the mutants of CYP102A1 are prepared by at least one selected from a group consisting of: substituting 47 th  amino acid arginine (R) of wild-type CYP102A1 with one amino acid selected from a group consisting of alanine, valine, leucine, isoleucine, proline, methionine, phenylalanine, and tryptophan, substituting 51 st  amino acid tyrosine(Y) of wild-type CYP102A1 with one amino acid selected from a group consisting of phenylalanine, alanine, valine, leucine, isoleucine, proline, methionine, tryptophan, substituting 64 th  amino acid glutamic acid (E) of wild-type CYP102A1 with one amino acid selected from a group consisting of glycine, serine, threonine, cysteine, tyrosine, asparagine, and glutamine, substituting 74 th  amino acid alanine (A) of wild-type CYP102A1 with one amino acid selected from a group consisting of glycine, serine, threonine, cysteine, tyrosine, asparagine, and glutamine, substituting 81 st  amino acid phenylalanine (F) of wild-type CYP102A1 with one amino acid selected from a group consisting of alanine, valine, leucine, isoleucine, proline, methionine, and tryptophan, substituting 86 th  amino acid leucine (L) of wild-type CYP102A1 with one amino acid selected from a group consisting of alanine, valine, isoleucine, proline, methionine, phenylalanine, and tryptophan, substituting 87 th  amino acid phenylalanine (F) of wild-type CYP102A1 with one amino acid selected from a group consisting of alanine, valine, leucine, isoleucine, proline, methionine, and tryptophan, substituting 143 rd  amino acid glutamic acid (E) of wild-type CYP102A1 with one amino acid selected from a group consisting of glycine, serine, threonine, cysteine, tyrosine, asparagine, and glutamine, substituting 188 th  amino acid leucine (L) of wild-type CYP102A1 with one amino acid selected from a group consisting of glycine, serine, threonine, cysteine, tyrosine, asparagine, and glutamine, and substituting 267 th  amino acid glutamic acid (E) of wild-type CYP102A1 with one amino acid selected from a group consisting of alanine, valine, leucine, isoleucine, proline, methionine, phenylalanine, and tryptophan. 
     
     
         11 . The method of  claim 6 , wherein the mutants of CYP102A1 are prepared by at least one selected from a group consisting of: substituting 47 th  amino acid arginine (R) of wild-type CYP102A1 with leucine (L), substituting 51 st  amino acid tyrosine(Y) of wild-type CYP102A1 with phenylalanine, substituting 64 th  amino acid glutamic acid (E) of wild-type CYP102A1 with glycine (G), substituting 74 th  amino acid alanine (A) of wild-type CYP102A1 with glycine (G), substituting 81 st  amino acid phenylalanine (F) of wild-type CYP102A1 with isoleucine (I), substituting 86 th  amino acid leucine (L) of wild-type CYP102A1 with isoleucine (I), substituting 87 th  amino acid phenylalanine (F) of wild-type CYP102A1 with valine (V), substituting 143 rd  amino acid glutamic acid (E) of wild-type CYP102A1 with glycine (G), substituting 188 th  amino acid leucine (L) of wild-type CYP102A1 with glutamine (Q), and substituting 267 th  amino acid glutamic acid (E) of wild-type CYP102A1 with valine (V). 
     
     
         12 . The method of  claim 6 , wherein the mutants of CYP102A1 comprises amino acid substitution sites of wild-type CYP102A1 selected from a group consisting of F87A, R47L/Y51 F, A74G/F87V/L188Q, R47L/L86I/L188Q, R47L/F87V/L188Q, R47L/F87V/L188Q/E267V, R47L/L86I/L188Q/E267V, R47L/L86I/F87V/L188Q, R47L/F87V/E143G/L188Q/E267V, R47L/E64G/F87V/E143G/L188Q/E267V, R47L/F81I/F87V/E143G/L188Q/E267V, and R47L/E64G/F81I/F87V/E143G/L188Q/E267V. 
     
     
         13 . A kit for preparing piceatannol from resveratrol, the kit comprising at least one enzyme selected from a group consisting of wild-type CYP102A1 and mutants of CYP102A1, and an NADPH-generating system. 
     
     
         14 . The kit of  claim 13 , wherein the NADPH-generating system comprises glucose 6-phosphate, NADP+, and yeast glucose 6-phosphate.