Functional properties proteases

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. 

Kuehler and Stine (43) studied the functional properties of whey protein with respect to emulsifying capacity as affected by treatment with three proteolytic enzymes. Two microbial proteases and pepsin were examined. The emulsion capacity decreased as proteolysis continued, suggesting that there is an optimum mean molecular size of the whey proteins contributing to emulsification. 

The expanded use of enzymes to modify protein functional properties has great promise for the food industry. Major advantages of using proteases compared to other agents include their specificity, their effectiveness at low concentrations and... 

Existing uses of proteases in foods have been discussed in the foregoing section. Expanding such applications in the future depends upon our ability to control both the processes themselves and their costs. The development of continuous reactors utilizing free or immobilized enzymes will address each of these constraints. Furthermore, our understanding of the chemical basis for the various functional properties of proteins must be expanded... 

That is, we must learn how amino acid content and molecular configuration of food proteins are related to their functional properties. This goal is made more difficult by the fact that secondary, tertiary, and quarternary structures of proteins are likely to be quite different when exerting functional effects in food systems as compared to structures of the same proteins in dilute solutions and in their native states. The way in which specific actions of proteases affect protein structure must also be studied so that correlations with changes in functional properties can be made (61). 

In this context, Greene and coworkers [159, 160] have reported the first low-molecular-mass immunoglobulin mimetic 207, Scheme 62, developed on the basis of an X-ray structure analysis of the antigen-antibody complex. Compound 207 is resistant toward proteases and imitates the binding and functional properties of the native antibody. 

More details of the plastein reaction and its application to remove pigments such as chlorophyll, or to remove off-flavor components such as the beany taste of soybeans, are shown in Figure 2. The protein of the food system is solubilized and denatured (in order to achieve proteolysis), a protease is added, and the hydrolytic reaction is allowed to proceed. On partial hydrolysis of the protein the pigments and flavor constituents are released from the protein they are removed, the hydrolyzate is concentrated, and resynthesis and/or rearrangement of the amino acid sequence of the polypeptides is catalyzed by the same or a different protease. Resynthesis also can be carried out in the presence of added amino acid esters in order to improve the nutritional/functional properties of the protein. 

Kristinsson, H. G., and Rasco, B. A. 2000a. Biochemical and functional properties of Atlantic salmon (salmo salar) muscle proteins hydrolyzed with various alkaline proteases. /. Agric. Food Chem., 48, 657-666. 

Babiker, E.F.E., Fujisawa, N., Matsudomi, N., Kato, A. (1996). Improvement of the functional properties of gluten by protease digestion or acid hydrolysis followed by microbial transglutaminase treatment. J. Agric. Food Chem., 44, 3746-3750. 

Caessens, PJ.W.R. et al., P-Lactoglobulin hydrolysis. 1. Peptide composition and functional properties of hydrolysates obtained by the action of plasmin, trypsin, and Staphylococcus aureus V9 protease, J. Agric. Food Chem., 47, 2973, 1999. 

Chirality has had a profound effect on the structural and functional properties of biomolecules. For example, the right-handed helices observed in proteins result from the exclusive presence of L-amino acids. Polypeptides synthesized in the laboratory from both D- and L- amino acids do not form helices. In addition, because the enzymes are chiral molecules, they only bind substrate molecules in one enantiomeric form. Proteases, enzymes that degrade proteins by hydrolyzing peptide bonds, cannot degrade artificial polypeptides composed of D-amino acids. 

The interaction between proteases and their inhibitors is, at least by comparison with interactions which take place in food systems, a remarkably simple and straightforward association. Simple correlations of functional properties with different kinds of molecular forces cannot be made. It is possible, however, to illustrate the importance of protein structure and of protein—protein interactions as determinants of the functional properties of food proteins. I would like therefore to look at several food systems in which protein—protein or protein—other constituent interactions play a role and examine the relationship between functionality and protein structure in these systems. One of the simplest areas in which to examine this relationship is the well studied collagen-gelatin transition. 

One of the current approaches to the improvement of the functional properties of proteins is enzymatic hydrolysis [148], The emulsifying ability of soy protein isolate can be increased by treatment with neutral fungal protease however, this treatment decreases emulsion stability [163], Partial hydrolysis of fish protein concentrate improves both emulsification and stability [164]. On the other hand, treatment of whey protein concentrate with pepsin, pronase, and pro-lase leads to a decrease in emulsification ability, suggesting that there... 

Kato et al. [54,55] reported using the proteases papain, pronase E, and chymotrypsin for the deamidation of food proteins, and Kato et al. [56] later reported deamidation of selected plant and animal proteins with chymotrypsin immobilized on controlled pore glass. The reactions were carried out at alkaline pH (pH 10) and 20°C and resulted in 5 to 20% deamidation with up to 8% peptide bond hydrolysis. Functional properties of proteins thus deamidated showed increased solubility and emulsifying and foaming properties. Enzymatic... 

Gallagher et al. [138] investigated the future application of two enzymes, bromelain and a bacillus protease (Bacillus subtilis) in the production of peptides from casein in point of view of the functional properties of the products. Bromelain action resulted in a hydrolysate with a great number of high-molecular-mass peptides this may have improved the functional properties of a food product. The bacillus protease seemed to be more suitable for producing bitter peptides for future research and/or for future food. 

Deamidation of proteins is important for improving functional properties of the product under mild reaction conditions. But enzymatic deamidation of proteins has not had real attention until recently. Kato et al. [155] developed a method for enzymatic deamidation of food proteins by treatment with proteases at pH 10. Salt and disulfide reducing agents have little effect on soy protein deamidation. Heat treatment and proteolysis of soy proteins are the major factors affecting deamidation [156]. 

Hrckova, M. M. Rusnakova J. Zemanovic. Enzymatic hydrolysis of defatted soy flour by three different proteases and their effect on the functional properties of resulting protein hydrolysates. Czech. J. Food Sci. 2002,20, 7-14. 

Jung, S. RA. Murphy L.A. Johnson. Physicochemical and functional properties of soy protein substrates modified by low levels of protease hydrolysis./. Food Sci. 2005, 70, C180-187. 

Partial proteolysis of soy protein isolate with neutral protease from Aspergillus oryzae altered certain functional properties ( ). Solubility was increased in the enzyme-treated soy isolate at both neutral pH and at the isoelectric point (pH... 

Industrial applications represent more than 80% of the global market of enzymes. A distinction should be made between those cases in which the enzymatic conversion of the raw material into the product is the key operation and those in which the enzyme is used as an additive to modify certain functional property of the product. In the first case the enzymatic reaction is carried out in a controlled environment at optimized conditions with respect to the catalytic potential of the enzyme, while in the second case conditions for enzyme action are not specified to optimize its activity and sometimes not even controlled. Examples of the first case are the production of high-fructose syrups with immobilized glucose isomerase and the production of 6-aminopenicillanic acid from penicillin G with immobilized penicillin acylase examples of the second case are the use of fungal proteases in dough making and the use of pancreatin in leather bating. Most conventional uses of enzymes refer to... 

Photoinhibition at 20 C caused a significant loss of atrazine binding sites. This degradation of D1-protein was strongly inhibited at 0 C compared to 20 C (Fig.l). probably due to the inhibition of a membrane bound protease activity through lowering the temperature. The functional properties of reaction centre II. i.e. PSII-photochemistry (H2O —> SiMo)... 

Because the size of polypeptides produced from proteolysis relate directly to their functional properties, the application of DH is the most common approach to monitor the process of proteolysis. However, it should be pointed out that the calculation of PCL from the DH is a theoretical concept only. Although it has been widely used and documented in almost all of the literature, it cannot be used for comparing protein hydrolysates from different proteolytic reaction systems. When a protein substrate is hydrolyzed by different proteases, the differences in the specificity of enzymes may produce protein hydrolysate with significantly different-size distributions, although they have the same DH values. 

Heparin has been fluorescently labeUed in a manner that does not alter the functional properties of the polysaccharide. The labelled polysaccharide has been used in conjunction with fluorescence polarization spectroscopy to monitor the binding of heparin to the L-serine proteases thrombin, factor XIa, factor Xa, and plasmin. The stoicheiometry and dissociation constants of the interactions have been measured. The kinetics of inactivation of the four proteases by anti-thrombin as a function of heparin concentration have also been measured. Evidence shows that the direct binding of heparin to anti-thrombin is probably responsible for the polysaccharide-dependent acceleration of hemostatic enzyme-inhibitor reactions. 

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