Directed evolution of CYP52A12 gene and its use in dicarboxylic acid production
The invention relates to the directed evolution of CYP52A12 gene and the use thereof for the production of a dicarboxylic acid. In particular, it relates to a method of preparing a long chain dicarboxylic acid producing strain by using directed evolution and homologous recombination, a strain obtained by the method that is capable of producing a long chain dicarboxylic acid under an acidic condition and the use thereof. In particular, the invention relates to a method of preparing a long chain dicarboxylic acid producing strain by directed evolution of CYP52A12 gene and homologous recombination, a strain obtained by the method that is capable of producing a long chain dicarboxylic acid under an acidic condition and the use thereof. By directed evolution of CYP52A12 gene, one strain which has a base mutation at the promoter region of said gene and is capable of producing a long chain dicarboxylic acid under an acidic condition in a shortened fermentation time is screened out in the invention.
1. A product comprising an isolated modified CYP52A12 protein, wherein, with reference to the amino acid numbering of SEQ ID NO:20, the modified CYP52A12 protein comprises Asp at a position corresponding to the position 76 of SEQ ID NO: 20 and Ile at a position corresponding to the position 470 of SEQ ID NO: 20, compared to an unmodified CYP52A12 protein, and wherein the modified CYP52A12 protein comprises an amino acid sequence at least 99% identical to SEQ ID NO. 20 and has the capability of producing a long chain dicarboxylic acid under an acidic culture condition that is at least 95% or more of that of the modified CYP52A12 protein comprising the amino acid sequence set forth in SEQ ID NO.21.
2. The product of claim 1 , wherein the modified CYP52A12 protein comprises the amino acid sequence set forth in SEQ ID NO:21.