Increasing the catalytic power of the flavin reductase DszD enzyme using site-directed mutagenesis method in Rhodococcus erythropolis

The flavin reductase DszD enzyme is a key enzyme for providing required reduction potential in the bacterial desulfurization process. Considering the low speed of desulfurization process because of low catalytic power of this enzyme, it is necessary to increase the catalytic power of flavin reductase for industrial use of this enzyme as biocatalyst.

The three-dimensional structure of the flavin reductase DszD enzyme was predicted by a CPHmodel server and its amino acid sequence was searched in the protein data bank to identify the homologue molecules. Based on the alignment of the amino acid sequence and the model molecules, the key residues at the flavin mononucleotide substrate were identified. The key residue of asparagine at position 77 was replaced with phenylalanine using the site-directed mutagenesis method. 

 This study with research ethics code IR.NIGEB.EC.1398.6.24 A has been approved by research ethics committee at National Institute of Genetic Engineering and Biotechnology, Tehran, Iran.

 The cloning and expression of each of the wild-type and mutant genes were performed separately. The catalytic power of the produced wild-type and mutant enzymes were compared. The catalytic activity measurements showed that the mutant enzyme had a 2.5 fold increase in catalytic power.

 Replacing phenylalanine with asparagine at position 77 of flavin reductase DszD enzyme leads to an increase in enzyme catalytic power to increase the speed of bacterial desulfurization process.