Engineering methionine γ-lyase from Citrobacter freundii for anticancer activity

Publication date: Available online 27 September 2018Source: Biochimica et Biophysica Acta (BBA) - Proteins and ProteomicsAuthor(s): Samanta Raboni, Svetlana Revtovich, Nicola Demitri, Barbara Giabbai, Paola Storici, Chiara Cocconcelli, Serena Faggiano, Elena Rosini, Loredano Pollegioni, Serena Galati, Annamaria Buschini, Elena Morozova, Vitalia Kulikova, Alexey Nikulin, Edi Gabellieri, Patrizia Cioni, Tatyana Demidkina, Andrea MozzarelliAbstractMethionine deprivation of cancer cells, which are deficient in methionine biosynthesis, has been envisioned as a therapeutic strategy to reduce cancer cell viability. Methionine γ-lyase (MGL), an enzyme that degrades methionine, has been exploited to selectively remove the amino acid from cancer cell environment. In order to increase MGL catalytic activity, we performed sequence and structure conservation analysis of MGLs from various microorganisms. Whereas most of the residues in the active site and at the dimer interface were found to be conserved, residues located in the C-terminal flexible loop, forming a wall of the active site entry channel were found to be variable. Therefore, we carried out site-saturation mutagenesis at four independent positions of the C-terminal flexible loop, P357, V358, P360 and A366 of MGL from Citrobacter freundii, generating libraries that were screened for activity. Among the active variants, V358Y exhibits a 1.9-fold increase in the catalytic rate and a 3-fold increase in KM, resulting in a catalyti...
Source: Biochimica et Biophysica Acta (BBA) Proteins and Proteomics - Category: Biochemistry Source Type: research