Purification and Partial Characterization of a Detergent and Oxidizing Agent Stable Alkaline Protease from a Newly Isolated Bacillus subtilis VSG-4 of Tropical Soil

• Published in: The journal of microbiology Vol. 49; no. 3; pp. 455 - 461
• Main Authors: Giri, Sib Sankar, Periyar Maniammai University, Tamilnadu, India, Sukumaran, V., Periyar Maniammai University, Tamilnadu, India, Sen, Shib Sankar, Jadavpur University, Kolkata, India, Oviya, M., Periyar Maniammai University, Tamilnadu, India, Nazeema Banu, B., Periyar Maniammai University, Tamilnadu, India, Jena, Prasant Kumar, Nirma University, Gujarat, India
• Format: Journal Article
• Language: English
• Published: Heidelberg The Microbiological Society of Korea 01-06-2011; Springer Nature B.V; 한국미생물학회
• Subjects: alkaline protease; Ammonium; Ammonium sulfate; BACILLUS SUBTILIS; Bacillus subtilis - drug effects; Bacillus subtilis - enzymology; Bacillus subtilis - genetics; Bacillus subtilis - isolation & purification; Bacterial Proteins - chemistry; Bacterial Proteins - genetics; Bacterial Proteins - isolation & purification; Bacterial Proteins - metabolism; Biochemical characteristics; Biochemistry; Biomedical and Life Sciences; characterization; Chromatography; detergent stability; Detergents; Detergents - pharmacology; Dialysis; Endopeptidases - chemistry; Endopeptidases - genetics; Endopeptidases - isolation & purification; Endopeptidases - metabolism; Enzyme Stability; Enzymes; Hydrogen peroxide; Hydrogen-Ion Concentration; Laundry; Life Sciences; Microbiology; Molecular Sequence Data; Oxidants - pharmacology; Oxidizing agents; pH effects; Precipitation; PURIFICACION; PURIFICATION; Sequence Analysis, DNA; Serine proteinase; Sodium; Sodium lauryl sulfate; Soil; Soil Microbiology; Stains; Surfactants; Temperature; Temperature effects; Thermal stability; Tropical Climate; Tropical environments; 생물학; Tamil Nadu India; India; alkaline protease; characterization; detergent stability; purification
• ISSN: 1225-8873; 1976-3794
• DOI: 10.1007/s12275-011-0427-4

An extracellular detergent tolerant protease producing strain VSG-4 was isolated from tropical soil sample and identified as Bacillus subtilis based on morphological, biochemical characteristics as well as 16S-rRNA gene sequencing. The VSG-4 protease was purified to homogeneity using ammonium sulphate precipitation, dialysis and sephadex G-200 gel permeation chromatography with a 17.4 purification fold. The purified enzyme was active and stable over a broad range of pH (8.0-11.0, optimum at 9.0) and temperature (40℃ to 60℃, optimum at 50℃). The thermostability of the enzyme was significantly increased by the addition CaCl₂. This enzyme was strongly inhibited by PMSF and DFP, suggesting that it belongs to the serine protease superfamily. The purified VSG-4 alkaline protease showed remarkable stability in anionic (5 mM SDS) and ionic (1% Trion X-100 and 1% Tween 80) detergents. It retained 97±2% and 83.6±1.1% of its initial activity after 1 h preincubation in the presence of 1% H₂O₂ and 1% sodium perborate, respectively. Furthermore, the purified enzyme showed excellent stability and compatibility with some commercial laundry detergents besides its stain removal capacity. Considering these promising properties, VSG-4 protease may find tremendous application in laundry detergent formulations.