Effects in cancer cells of the recombinant L-Methionine gamma-lyase from Brevibacterium aurantiacum. Encapsulation in human erythrocytes for sustained L-methionine elimination

Methioninedeprivation induces growth arrest and death of cancer cells. To eliminateL- methioninewe produced, purified and characterized the recombinant pyridoxal59-phosphate (PLP)-dependent L- methionine gamma-lyaseMGL-BL929 from the cheese-ripening Brevibacteriumaurantiacum. Transformation of an Escherichia coli strain with the gene BL929 from B. aurantiacum optimized for E. coli expression led to production of the MGL-BL929. Eliminationof L- methionineand cytotoxicity in vitro were assessed, and methylation-sensitive epigenetics was explored for changes resulting from exposure of cancer cells to the enzyme. A bioreactor was built by encapsulation of the protein in human erythrocytes to achieve sustained eliminationof L- methioninein extracellular fluids. Catalysis was limited to α,γ- eliminationof L- methionineand L-homocysteine. The enzyme had no activity on other sulfur-containing amino acids. Enzyme activity decreased in presence of serum albumin or plasma resulting from reduction of PLP availability. Eliminationof L- methionineinduced cytotoxicity on a vast panel of human cancer cell lines and spared normal cells. Exposure of colorectal carcinoma cells to the MGL-BL929 reduced methyl-CpG levels of hyper methylated gene promoters including that of CDKN2Awhose mRNA expression was increased, together with decrease in global histone H3-dimethyl lysine9. The MGL erythrocyte bioreactor durably preserves enzyme activity in vitro and strongly eliminatesL- methioninefrom medium.