A novel acidic and SDS tolerant halophilic lipase from moderate halophile Nesterenkonia sp. strain F: molecular cloning, structure analysis and biochemical characterization

Abstract

A novel gene encoding an acidic and SDS tolerant halophilic lipase from moderately halophilic bacterium, Nesterenkonia sp. strain F (lipF) was cloned in Escherichia coli and the recombinant lipase (LipF) was characterized. The gene had 819 bp and encoded a protein of 272 amino acids residues (pI=7.0). No sequence similarity between LipF and other halophilic lipolytic enzymes was observed. LipF was grouped into the lipase family XIII because it contained a highly conserved motif GISMG. In contrast to halophilic proteins, equal amount of acidic and basic residues were observed in LipF. Also, a small amount of acidic residues was demonstrated on the surface of the LipF structure. After purification of the recombinant protein, it showed 28.5 kDa on SDS-PAGE. The enzyme was active over a pH range of 3.5 to 8.0 with optimum activity at pH 5.0. The optimum temperature was 55 °C and it was active at 25 to 85 °C. Although LipF was active at 0-4 M of NaCl and KCl, increasing activity was observed with increasing concentration of the salts and maximum activity was at 4 M. LipF was stimulated by Ca²⁺, Cu²⁺ and Al³⁺ but inhibited with Ni²⁺, PMSF and EDTA. While 50% of the activity was remained in the presence of 0.1% SDS, 0.5% Tween 20 and Triton X-100, 1% SDS increased the activity to 120%. These characteristics indicate wide potential applications of the halophilic lipase in industries. This is the first cloning and characterization of an acidic halophilic lipase from a moderately halophilic bacterium.