Activity Improvement and Vital Amino Acid Identification on the Marine-Derived Quorum Quenching Enzyme MomL by Protein Engineering.

MomL is a marine-derived quorum-quenching (QQ) lactonase which can degrade various N -acyl homoserine lactones (AHLs). Intentional modification of MomL may lead to a highly efficient QQ enzyme with broad application potential. In this study, we used a rapid and efficient method combining error-prone polymerase chain reaction (epPCR), high-throughput screening and site-directed mutagenesis to identify highly active MomL mutants. In this way, we obtained two candidate mutants, MomL _I144V and MomL _V149A . These two mutants exhibited enhanced activities and blocked the production of pathogenic factors of Pectobacterium carotovorum subsp. carotovorum (Pcc) . Besides, seven amino acids which are vital for MomL enzyme activity were identified. Substitutions of these amino acids (E238G/K205E/L254R) in MomL led to almost complete loss of its QQ activity. We then tested the effect of MomL and its mutants on Pcc -infected Chinese cabbage. The results indicated that MomL and its mutants (MomL _L254R , MomL _I144V , MomL _V149A ) significantly decreased the pathogenicity of Pcc . This study provides an efficient method for QQ enzyme modification and gives us new clues for further investigation on the catalytic mechanism of QQ lactonase.