Enzymatic production of l-citrulline by hydrolysis of the guanidinium group of l-arginine with recombinant arginine deiminase

In this study, a simple, efficient enzymatic production process for the environmentally friendly synthesis of l-citrulline from l-arginine was developed using arginine deiminase (ADI) from Lactococcus lactis. Following overexpression of L. lactis ADI in Escherichia. coli BL21 (DE3) and experimental evolution using error-prone PCR, mutant FMME106 was obtained with a Km for l-arginine of 3.5 mM and a specific activity of 195.7 U/mg. This mutant exhibited a maximal conversion of 92.6% and achieved a final l-citrulline concentration of 176.9 g/L under optimal conditions (190 g/L l-arginine, 15 g/L whole-cell biocatalyst treated with 2% isopropanol for 30 min, 50 °C, pH 7.2, 8 h). The average l-citrulline synthesis rate of 22.1 g/L/h is considerably higher than that reported for other similar biocatalytic approaches, therefore the process developed in the present work has great potential for large-scale production of l-citrulline.