Purification and biochemical characteristics of the protease from Lactobacillus brevis R4 isolated from Harbin dry sausages

Abstract

This study investigated the purification and biochemical characteristics of the protease secreted by Lactobacillus brevis R4 that was isolated from Harbin dry sausages. The optimized fermentation conditions were a fermentation time of 36 h, an initial pH of 5 and a fermentation temperature of 42 °C. A 27.9 kDa extracellular protease was purified using ammonium sulfate precipitation, ion exchange layer and gel filtration and was determined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). The protease produced by L. brevis R4 reached the highest relative protease activity at pH 7, 35 °C and 36 h. The microbial protease activity was inhibited by Zn2+, Cu2+ ions and ethylene diamine tetraacetic acid disodium salt (EDTA). The maximum velocity of the reaction (Vmax) and Michaelis constant (Km) of the protease were 65.9 mg/min and 17.1 mg/mL, respectively. SDS-PAGE demonstrated the ability of the protease to hydrolyse myofibrillar and sarcoplasmic proteins, especially on the myosin heavy chain, paramyosin, troponin, myosin light chain and glyceraldehyde dehydrogenase. The study provides a basis for understanding the enzymatic properties of the L. brevis R4 protease. In conclusion, L. brevis R4 has the potential to be used as a starter culture or protease-producing strain to produce Harbin dry sausages.