Profiles of Proteinases in Gouda-Type Cheese

The pH of buffers used for extraction of proteinases from Gouda-type cheese affected the kinds of extractable proteinases. Proteinases in the fractions extracted with pH 3.0, 4.0, and 6.0 buffers were separated by CM-Sephadex or diethylaminoethyl-cellulose. Two (F3-1 and F3-II) , three (F4-1, F4-II, and F4-III), and one (F6) proteinases were separated from the pH 3.0, 4.0, and 6.0 extracts. The F3-I was completely inhibited by pepstatin. The F3-I degradated asl-and /3-casein into products with the same mobilities as asl-casein(f24 to 199) and ]3-casein(f1 to 189) peptide, respectively. The F3-II was strongly inhibited by diisopropyl-fluorophosphate and soybean trypsin inhibitor, whereas F3-II produced fragments with mobilities equal to those of 3'-caseins, that is, j3-casein(f29 to 209), ~-casein(fl06 to 209), and H-casein (f1~9 to 209). The F4-I was completely inhibited by pepstatin as well as F3-1, but changes in the polyacrylamide gel electrophoresis pattern of casein by F4-I were different from those by F3-I. Properties of F 4-II and F 4-III were similar to those of F~-I and F3-1I; therefore, F4-1I and F4-1II are considered to be the same enzymes as F3-1 and F3-II, respectively. The F 6 was strongly inhibited by diisopropyl-fluorophosphate and ethylenediaminetetraacetic acid. In Gouda-type cheese, there are at least four proteinases, two of them being serine proteinases and the other two acid proteinases.