The use of bacteria with overexpression of the cadA gene for the bioconversion of lysine into cadaverine

Abstract

The article is devoted to construction of an Escherichia coli strain with overexpression of the cadA gene encoding lysine decarboxylase, as well as to assessment of its ability to convert lysine to cadaverine depending on the pH of the medium. The overexpressing strain was constructed on the base of E. coli BL21DE3 strain transformed with the pET19b plasmid carrying the cadA gene from E. coli MC4100. Bacteria were grown in 5 mL of LB broth at 37 °C without stirring, overexpression of cadA was induced by the addition of 1 mM isopropyl-β-D-thiogalactopyranoside (at OD600 = 0.6), after 2 hours the cells were washed and transferred to a medium with pH 7.4 or 4.0 with the addition 5 g/L L-lysine hydrochloride. The amount of synthesized cadaverine was quantified by TLC with preliminary derivatization with dansyl chloride. The maximum rate of bioconversion was observed when cells overexpressing cadA were cultivated in a medium with pH 4.0 (2.8 ± 0.7 mmol cadaverine/g dry weight per hour). The rate of bioconversion under basal conditions (pH 7.4, basal expression) was 6.8 times lower. The final concentration of cadaverine when cultivated in a medium with pH 7.4 and/or in the absence of overexpression of the cadA gene was 0.8-1 mM; under conditions of overexpression of the gene, up to 1.2 mM of cadaverine accumulated in a neutral medium, and 1.8 mM in an acidic medium.