Purification and Characterization of an Alkaline Protease from Micrococcus sp. Isolated from the South China Sea

• Published in: Journal of Ocean University of China Vol. 16; no. 2; pp. 319 - 325
• Main Authors: Hou, Enling, Xia, Tao, Zhang, Zhaohui, Mao, Xiangzhao
• Format: Journal Article
• Language: English
• Published: Heidelberg Science Press 01.04.2017; Springer Nature B.V
• Subjects: Ammonium; Bacteria; Biotechnology; characterization; China; Chromatography; Cosmetics; Earth and Environmental Science; Earth Sciences; Enzymatic activity; Ionization; Marine; Meteorology; Micrococcus; Oceanography; protease; Proteases; purification; Sea; South; the; South China Sea; ISEW, South China Sea; protease; characterization; sp; purification; the South China Sea
• ISSN: 1672-5182; 1993-5021; 1672-5174
• DOI: 10.1007/s11802-017-3207-x

Protease is wildly used in various fields, such as food, medicine, washing, leather, cosmetics and other industrial fields.In this study, an alkaline protease secreted by Micrococcus NH54PC02 isolated from the South China Sea was purified and characterized.The growth curve and enzyme activity curve indicated that the cell reached a maximum concentration at the 30th hour and the enzyme activity reached the maximum value at the 36th hour. The protease was purified with 3 steps involving ammonium sulfate precipitation, ion-exchange chromatography and hydrophobic chromatography with 8.22-fold increase in specific activity and23.68% increase in the recovery. The molecular mass of the protease was estimated to be 25 kDa by SDS-PAGE analysis. The optimum temperature and pH for the protease activity were 50℃ and pH 10.0, respectively. The protease showed a strong stability in a wide range of pH values ranging from 6.0–11.0, and maintained 90% enzyme activity in strong alkaline environment with pH 11.0.Inhibitor trials indicated that the protease might be serine protease. But it also possessed the characteristic of metalloprotease as it could be strongly inhibited by EDTA and strongly stimulated by Mn2+. Evaluation of matrix-assisted laser desorption ionization/time-of-flight MS (MALDI-TOF-TOF/MS) showed that the protease might belong to the peptidase S8 family.