Patent ID: 8895307

Claim:
A method for the evolution of a NADPH using ketol-acid reductoisomerase enzyme to an NADH using form comprising: (a) providing a ketol-acid reductoisomerase enzyme wherein said ketol-acid reductoisomerase enzyme has an amino acid sequence that matches the Profile Hidden Markov Model for KARIs in Table 9 with an E value of <10 −3 using the HMMER 2.2 g hmmsearch program in the HMMER 2.2 g package with the Z parameter set to 1 billion and uses NADPH as a cofactor; (b) identifying the cofactor switching residues in the enzyme of a) corresponding to the amino acids at positions 24, 33, 47, 50, 52, 53, 61, 80, 115, 156, 165, and 170 of SEQ ID NO: 17; (c) creating mutations in at least two of the cofactor switching residues of b) to create a mutant enzyme, wherein one of the at least two mutations is a substitution of the cofactor switching residue corresponding to position 52 of SEQ ID NO: 17.