Kinetic Parameters and Cytotoxic Activity of Recombinant Methionine γ-Lyase from Clostridium tetani, Clostridium sporogenes, Porphyromonas gingivalis and Citrobacter freundii

The steady-state kinetic parameters of pyridoxal 5'-phosphate-dependent recombinant methionine γ-lyase from three pathogenic bacteria, Clostridium tetani, Clostridium sporogenes, and Porphyromonas gingiva- lis, were determined in β- and γ-elimination reactions. The enzyme from C. sporogenes is characterized by the highest catalytic efficiency in the γ-elimination reaction of L-methionine. It was demonstrated that the enzyme from these three sources exists as a tetramer. The N-terminal poly-histidine fragment of three recombinant enzymes influences their catalytic activity and facilitates the aggregation of monomers to yield dimeric forms under denaturing conditions. The cytotoxicity of methionine γ-lyase from C. sporogenes and C. tetani in com- parison with Citrobacter freundii was evaluated using K562, PC-3, LnCap, MCF7, SKOV-3, and L5178y tumor cell lines. K562 (IC 50 =0.4-1.3 U/ml), PC-3 (IC 50 =0.1-0.4 U/ml), and MCF7 (IC 50 =0.04-3.2 U/ml) turned out to be the most sensitive cell lines. KEYWORDS kinetic parameters; methionine γ-lyase; pathogenic microorganisms; oligomeric structure; cytotox- icity. ABBREVIATIONS DTT - dithiothreitol; His-tag - poly-histidine fragment; MGL - methionine γ-lyase; MSC - mes- enchymal stromal cells; PLP - pyridoxal 5'-phosphate.