Purification and characterization of soymilk-clotting enzymes from Bacillus sp. K-295G-7.

Abstract

Enzymes I and II, which have a high soymilk-clotting activity, produced from K-295G-7 were purified by chromatographies on Sephadex G-100, CM-cellulose, hydroxylapatite, and 2nd Sephadex G-100. The two purified enzymes were found to be homogeneous by polyacrylamide gel electrophoresis (PAGE) at pH 4.3. The molecular weights ofenzymes I and II were 28,000 and 29,500 by SDS-PAGE, and their isoelectric points were 9.22 and 9.45, respectively. Enzymes I and II coagulated soymilk optimally at 65°C and were stable up to 45°C. Both enzymes were most active at pH 5.8, for soymilk coagulation between pH 5.8 to 6.7, and were stable with about 50~100% of the original activity from pH 5 to 10. Each of the purified enzymes was a serine protease with an optimum pH of9.0 for soy protein isolate (SPI) and casein digestions, because these enzymes were inhibited completely by diisopropylfluoro-phosphate (DFP). The soymilk-clotting activity to proteolytic activity ratio of the enzyme II was 3 times higher than that of enzyme I. Enzymes I and II were more sensitive to the calcium ion concentration in soymilk than bromelain is. Soybean contains useful human nutrients such as proteins, lipids and sugars. Soybean

Utilization of Salt Whey From Egyptian Ras (Cephalotyre) Cheese in Microbial Milk Clotting Enzymes Production

“…(2003) arrived to the same conclusion with M. pusillus protease which increased in its clotting activity the increase in Ca++ ion. Park et. al.…”

Identification of Strong Aggregating Regions in Soy Glycinin upon Enzymatic Hydrolysis

“…These proteases are subtilisin Carlsberg from Bacillus licheniformis – and proteases that originate from Bacillus sp. K-295G-7 – and Bacillus pumilus TYO-67 , . The plant proteases bromelain , and papain and the animal-derived protease chymotrypsin were also shown to induce aggregation of soy proteins.…”

Insights into the Reaction Mechanism of the Coagulation of Soy Protein Isolates Induced by Subtilisin Carlsberg

“…Proteins prepared from defatted soybean meal show coagulation by treatment with bromelain, and the coagulation develops with increasing amounts of the fragments formed from the basic subunit of an 11S globulin (13). Some microbial enzymes have a clotting activity of soybean milk (14,15). A serine protease of Bacillus pumilus shows the coagulating activity by degrading the acidic subunit of an 11S globulin and R′-, R-, and β-subunits of a 7S globulin but not the basic subunit of the 11S globulin (15).…”