Ca²⁺-induced stabilization of the nucleoside 2′- deoxyribosyltransferase from Lactobacillus hilgardii ZJS01: Characteristics and application in nucleosides synthesis

A novel nucleoside 2′-deoxyribosyltransferase (LhNDT) from Lactobacillus hilgardii ZJS01 was cloned and expressed in Escherichia coli BL21 (DE3). The amino acid sequence of LhNDT displayed the highest identity of 26% to other reported NDTs. LhNDT was a monomer as revealed by gel filtration chromatography with or without addition of Ca²⁺ in elution buffer. This enzyme showed a high activity and stability over a broad pH range and was sensitive to temperature. With the addition of 15 mM Ca²⁺, the thermostability of LhNDT was greatly promoted by 63 times and 46 times at 30 °C and 35 °C, respectively. With the use of 2′-deoxyuridine as sugar donor at an economic molar ratio of 1:1, 5-aza-2′-deoxycytidine (46%) and 5-fluoro-2′- deoxycytidine (58%) were efficiently synthesized from 5-azacytosine and 5-fluorocytosine, respectively, by LhNDT for 2 h.