Insights into the reaction mechanism of the coagulation of soy protein isolates induced by subtilisin carlsberg.
The reaction mechanism of the coagulation of soy protein isolates (SPIs) induced by subtilisin Carlsberg was investigated. Formation of the coagula was monitored by measuring the turbidity (OD660) of the SPI solution, which decreased at the initial stage (phase 1 or digestion phase) of the reaction, and then increased (phase 2 or coagulation phase) and finally reached the plateau level. The velocity of the coagulation increased with increasing enzyme concentration. The coagulation was inhibited dramatically by adding a serine protease inhibitor (phenylmethanesulfonyl fluoride, PMSF) when the turbidity reached the minimum value. This indicates that the SPI digests participating in the coagulation are produced mainly in phase 2; in other words, production of the coagulating fragments and their coagulation occur simultaneously in phase 2. Structural changes of SPI during proteolysis were measured by observing fluorescence changes of aromatic amino acids of SPI and an externally added hydrophobic probe. It was suggested that the hydrophilic surface areas of SPIs might be cleaved preferentially in phase 1, and that the hydrophobic inner areas might be cleaved in phase 2 with extensive decomposition of the 3-D structure of SPI proteins. The fragments formed in phase 2 are considered to coagulate through hydrophobic interactions.
Nagai
Kouhei
K
Division of Food Science and Biotechnology, Graduate School of Agriculture, Kyoto University, Sakyo-ku, Kyoto 606-8502, Japan.
Inouye
Kuniyo
K
D024322:Amino Acids, Aromatic / Q000737:chemistry; D004591:Electrophoresis, Polyacrylamide Gel; D009391:Nephelometry and Turbidimetry; D010664:Phenylmethylsulfonyl Fluoride / Q000494:pharmacology; D015842:Serine Proteinase Inhibitors / Q000494:pharmacology; D030262:Soybean Proteins / Q000737:chemistry* / Q000302:isolation & purification*; D013381:Subtilisins / Q000494:pharmacology*
D024322:Amino Acids, Aromatic; D015842:Serine Proteinase Inhibitors; D030262:Soybean Proteins; D010664:Phenylmethylsulfonyl Fluoride; D013381:Subtilisins