Method for preparing r-hydroxynitrile lyase and use thereof

A method for preparing R-hydroxynitrile lyase, comprising:

• mutating R-hydroxynitrile lyase wild-type gene, comprising:
• mutating the wild-type gene sequence of R-hydroxynitrile lyase set forth in SEQ ID NO. 1 to obtain a hydroxynitrile lyase mutant gene having the gene sequence set forth in SEQ ID NO. 2, wherein:
• bases ATC at positions 325-327 of SEQ ID NO. 1 are substituted for bases ATG,
• bases AAT at positions 328-330 of SEQ ID NO. 1 are substituted for bases AGT,
• bases ATT at positions 961-963 of SEQ ID NO. 1 are substituted for bases ACT,
• bases CCA at positions 1060-1062 of SEQ ID NO. 1 are substituted for bases ATA, and
• bases TCT at positions 1063-1065 of SEQ ID NO. 1 are substituted for bases TTA;
• adding an enzyme cleavage site, comprising:
• adding a double digestion site to the hydroxynitrile lyase mutant gene, wherein the double digestion site is Ndel/Hindllll;
• preparing a recombinant plasmid, comprising:
• inserting the hydroxynitrile lyase mutant gene into an expression vector to obtain a recombinant plasmid, wherein the expression vector is pET26b(+), N-terminus of the expression vector is a signal peptide pelB leader having the gene sequence set forth in SEQ ID NO. 5;
• introducing the recombinant plasmid into a strain, comprising:
• introducing the recombinant plasmid with the hydroxynitrile lyase mutant gene into the strain to obtain a recombinant expression strain, wherein the strain is E. coli BL21 (DE3);
• and
• expressing and secreting the R-hydroxynitrile lyase in the strain, comprising:
• culturing the recombinant expression strain, and inducing the recombinant expression strain to express the enzyme of R-hydroxynitrile lyase in a culture medium and collecting a resulting enzyme liquid.