Purification and characterization of a cell surface-associated esterase from Lactobacillus fermentum DT41

A cell surface-associated esterase from Lactobacillus fermentum DT41, a starter used to produce Parmesan cheese, was purified to homogeneity by chromatography on Q-Sepharose, Sephacryl 200, Phenyl-Superose and Mono Q. The enzyme, together with a distinct cytoplasmic esterase, expressed the highest activity during the exponential phase of growth. The esterase was a monomer with a M r of ca 67 kDa and was most active at pH 7.0 and 30–35 °C, retaining considerable activity at pH 5.0 and 15 °C. The enzyme was stable at the cooking temperature (54–56 °C for a few minutes) of Parmesan cheese, its D 55 °C value was 35 min. The highest activity was determined on β-naphthyl butyrate, but marked hydrolysis also occurred with β-naphthyl esters of C2 to C10 fatty acids. β-Naphthyl esters of C14 to C18:1 fatty acids were not hydrolyzed and only tributyrin was degraded among the triglycerides. The K m on β-naphthyl butyrate was 0.31 mM with a V max of 140 μmol min −1mg −1. The esterase was strongly inhibited by 5 mM phenylmethylsulfonyl fluoride and by 1 mM Hg 2+ and Ag +, and moderately stimulated by Ca 2+ and Mg 2+.