Characterization of a novel carbohydrate esterase 7 family acetyl xylan esterase from Thermobifida halotolerans YIM 90462T

Abstract

The newly semi-synthetic β-lactam antibiotics play critical roles in prevention and treatment of β-lactam antibiotic-resistant bacteria. Acetyl xylan esterases belong to carbohydrate esterase 7 family can specifically hydrolyze 7-aminocephalosporanic acid (7-ACA) to produce deacetyl-7-aminocephalosporanic acid (D-7ACA) which is one of the most important starting materials to synthesize newly semi-synthetic β-lactam antibiotics. Here, an effective acetyl xylan esterase, ThAXE, from Thermobifida halotolerans YIM 90462 T, was firstly identified and characterized. ThAXE exhibited its highest catalytic activity against p-nitrophenyl acetate (p-NPC2) at pH 8.5 and 50 °C, with Km and Kcat/Km were 1.0 ± 0.1 mM and 2262.9 mM−1 s−1, respectively. Additionally, ThAXE can hydrolyze the acetyl group of 7-ACA at its C3' position with the specific activity of 7.4 U mg−1. Furthermore, molecular docking combined with site-directed mutagenesis validation demonstrated that the pocket of catalytic residues Ser186, Asp272, and His301 could accommodate p-NPC2 and 7-ACA ligands. This is the first report of a novel acetyl xylan esterase from T. halotolerans that has considerable potential biotechnological values.