Milk protease

Kaminogawa, S., Mizobuchi, H. and Yamauchi, K. 1972. Comparison of bovine milk protease with plasmin. Agr. Biol. Chem. 36, 2163-2167. 

The other major casein in cheese is /3-casein, but it is generally not hydrolyzed by rennet in low-pH cheeses. Alkaline milk protease (plas-min) plays the major role in the hydrolysis of /3-casein (Richardson and Pearce 1981). The plasmin level in cheese is related to the pH of the curd at whey drainage, since plasmin dissociates from casein micelles as the pH is decreased. Richardson and Pearce (1981) found two or three times more plasmin activity in Swiss cheese than in Cheddar cheese. Swiss cheese curds are drained at pH 6.4 or higher, while Cheddar cheese curds are drained at pH 6.3 or lower. Proteolysis of /3-casein is significantly inhibited by 5% sodium chloride. The inhibitory influence of sodium chloride is most likely due to alteration of /3-casein or a reduction in the attractive forces between enzyme and substrate (Fox and Walley 1971). 

Noomen, A. 1978. Activity of proteolytic enzymes in simulated soft cheeses (Meschanger type). 1. Activity of milk protease. Neth. Milk Dairy J. 32, 26-48. 

Kaminogawa et al. (1972) studied the milk protease, which is presumably responsible for the heterogeneity of galactosyltransferase, in comparison with plasmin, and they reported that these two enzymes may be the same. Subsequently, this proposal was confirmed (see Andrews, 1983). Powell and Brew (1974b) concluded that galactosyltransferase occurs in mature milk as a proteolytically degraded form of the galactosyltransferase that first appears in colostrum. 

Sainz et al. (2009) examined the combined effect of pressure (300-600MPa), temperature (40°C-60°C), and homogenization on the protease activity in milk. Inactivation of protease could extend the shelf life of milk. Protease was found to be very resistant to high pressures pressure stability was higher in raw milk than in pasteurized milk and... 

Milk is known to contain a protease indigenous to the casein micelle (203). Survival of this protease during the cheesemaking process would allow this proteolytic enzyme to exert its activity. Indeed it has been proposed that the milk protease plays a significant role in the aging of cheese (204), and the /3-casein is its principal substrate (204, 205, 206). 

Proteolysis has three phases (see Section 2.4.5.2.1) proteolysis in milk before cheese manufacture due to indigenous milk protease (plasmin) activity, the enzymatically induced coagulation of the milk in rennet cheeses (hydrolysis of K-casein by rennin) and proteolysis dining ripening of most cheeses, which is the most important reaction having a major impact on flavour and texture. 

The enzymatic hydrolysates of milk casein and soy protein sometimes have a strong bitter taste. The bitter taste is frequently developed by pepsin [9001 -75-6] chymotrypsin [9004-07-3] and some neutral proteases and accounted for by the existence of peptides that have a hydrophobic amino acid in the carboxyhc terminal (226). The relation between bitter taste and amino acid constitution has been discussed (227). 

Escherichia coli. The genetics of this gram-negative bacterium are very well known. For this reason, many of the first efforts to produce recombinant products from this microorganism were successful. However, because of the importance of the other criteria Hsted above, many efforts failed. E. co/i is only used to produce the milk-clotting mammalian protease chymosin [9001-98-3] (rennin). 

Two other practical appHcations of en2yme technology used in dairy industry are the modification of proteins with proteases to reduce possible allergens in cow milk products fed to infants, and the hydrolysis of milk with Hpases for the development of Hpolytic flavors in speciaHty cheeses. 

Inflammatory cells produce profeases, which allow the cells to enter the affected area. Some pathogens also produce proteases in order to enter the body. Human milk contains active protease inhibitors (e.g., ot-1-antitr) sin, a-l-antichymotr) sin, and elastase inhibitor) that can limit the ability of pafhogens fo gain enfry info fhe body and limit the inflammation caused by the inflammatory response (Lindberg et al., 1982). 

Lindberg, T., Ohlsson, K., and Westrom, B. (1982). Protease inhibitors and their relation to protease activity in human milk. Pediatr. Res. 16,479 83. 

The pretreatment of wastewater with hydrolases or acids is one of the best ways to overcome this obstacle, because in this way the big polymer molecules can be decomposed to smaller units, which can be measured by the biosensor. The positive effect of an enzymatic pretreatment of wastewater prior to sensorBOD measurement was demonstrated [53, 66]. In these investigations, different types of wastewaters, which contained milk powder, starch, or cellulose, were treated by proteases, a-amylases, and cellulases or a mix of these enzymes, respectively. This pretreatment resulted in a good correlation between sensorBOD and the conventional five-day BOD, while the sensorBOD values for untreated wastewater were significantly lower (see Table 6). As an example, the sensorBOD of a wastewater from a paper factory increased approximately to the fourfold value when treated by a mixture of cellulase and -glucosidase. 

Apphed biocatalysis has its roots in ancient China and Japan in the manufacture of food and alcohohc drinks. Without knowing, man utilized microbial amylases and proteases, in particrrlar for the production of soy-derived foods. In Etrrope too, applied biocatalysis has a long history. Cheese making has always involved the use of enzymes. As far back as about 400 BC, Homer s Iliad mentions the use of kid stomach for making cheese. It was discovered that milk, which was stored in a bag made of a stomach of a recently slaughtered calf, lamb or kid was converted into a semi-sohd substance. Upon pressing of this substance a drier material was obtained (namely cheese) which... 

Lab products were used to produce cheese (Knapp, 1847). Berzelius is cited with details stating that 1 part of lab ferment preparation (essentially proteases) coagulates 1800 parts of milk, and that only 0.06 parts of the lab ferment is lost. 

We have chosen to discuss enzyme modification of proteins in terms of changes in various functional properties. Another approach might have been to consider specific substrates for protease action such as meat and milk, legumes and cereals, and the novel sources of food protein such as leaves and microorganisms ( ). Alternatively, the proteases themselves provide categories for discussion, among which are their source (animals, plants, microorganisms), their type (serine-, sulfhydryl-, and metalloenzymes), and their specificity (endo- and exopeptidases, aromatic, aliphatic, or basic residue bond specificity). See Yamamoto (2) for a review of proteolytic enzymes important to functionality. 

Heat-Resistant Lipases. The heat-resistant lipases and proteinases and their effects on the quality of dairy products have been reviewed (Cogan 1977, 1980). Several reports have linked the lipases from bacteria with the off-flavor development of market milk (Richter 1981 Shipe et al. 1980A Barnard 1979B). The microflora developing in holding tanks at 4°C [and presumably in market milk stored at 40°F (Richter 1981)] may produce exocellular lipases and proteases that may survive ordinary pasteurization and sterilization temperatures. Rancidity of the cheese and gelation of UHT milk appear to be the major defects caused by the heat-resistant enzymes. 

Nathans, G. R. and Hade, E. P. K. 1978. Bovine milk xanthine oxidase. Purification by ultrafiltration and conventional methods which omit addition of proteases. Some criteria for homogeneity of native xanthine oxidase. Biochim. Biophys. Acta 526, 328-344. 

Several proteases from animal organs have been investigated for their milk-clotting potential, but only chymosin, porcine pepsin, and bovine pepsin are of interest to the cheese industry. 

Fungal proteases have been investigated extensively in search of suitable milk clotting enzymes. Patents have been issued for production of rennets from E. parasitica, M. Pusillus var. Lindt and M. miehei var. Cooney et Emerson. These have been approved in the United States as secondary direct food additives (FDA. 1984B) and have experienced considerable commercial success in the United States as milk-clotting enzymes for cheese manufacture. Many other fungal sources have also been tried in the effort to find an inexpensive replacement for chymosin. 

M. pusillus var. Lindt protease has given satisfactory results as a chymosin substitute in the manufacture of a number of cheese varieties, but not all varieties of M. pusillus var. Lindt are capable of producing acceptable cheese (Babel and Somkuti 1968). The clotting activity of M. pusillus var. Lindt protease is more sensitive to pH changes between 6.4 and 6.8 than chymosin, but is much less sensitive than that of porcine pepsin (Richardson et al 1967). The same authors reported that CaCL added to milk affected the clotting activity of M. pusillus var. Lindt rennet more than it did that of chymosin rennet. They also reported that this rennet was more stable than chymosin between pH 4.75 and 6.25. M. pusillus var. Lindt rennet is not destroyed during the manufacture of Cheddar cheese, although less than 2% of the enzyme added to the milk remains in the curd. Nearly all of it is found in the whey (Holmes et al. 1977). Mickelsen and Fish (1970) found M. pusillus var. Lindt rennet to be much less proteolytic than E. parasitica rennet but more proteolytic than chymosin rennet on whole casein, a8-casein and /3-casein at pH 6.65. 

Many enzymes extracted from higher plants have been tried for clotting cheese milk (Burnett 1976), however, attempts to use them have been unsuccessful. Most plant proteases are strongly proteolytic and cause extensive digestion of the curd, which has resulted in reduced yields, bitter flavors, and pasty-bodied cheese. 

Babel, F. J. and Somkuti, G. A. 1968. Mucor pusillus protease as a milk coagulant for cheese manufacture. J. Dairy Sci. 51, 937-937. 

Foltmann, B. 1981. Mammalian milk-clotting proteases Structure, function, evolution and development. Neth. Milk Dairy J. 35, 223-366. 

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