Chemistry: molecular biology and microbiology – Enzyme – proenzyme; compositions thereof; process for... – Oxidoreductase
Reexamination Certificate
2000-05-24
2002-07-16
Prats, Francisco (Department: 1651)
Chemistry: molecular biology and microbiology
Enzyme , proenzyme; compositions thereof; process for...
Oxidoreductase
C435S068100, C530S402000
Reexamination Certificate
active
06420148
ABSTRACT:
FIELD OF THE INVENTION
This invention relates to a novel method for cross-linking a protein using an enzyme. More particularly, it relates to a novel method for cross-linking protein using a multi-copper oxidase such as laccase or bilirubin oxidase.
BACKGROUND OF THE INVENTION
Protein materials which are cross-linked to have a high molecular weight or gelled by the cross-linking method of the present invention can be used in the field of food processing such as of raw fish meat paste, kamaboko (fish cake), fish/livestock meat sausage, tofu (soy bean curd), noodles, confectionery/bread, food adhesives, sheet-like meat food, yogurt, jelly and cheese. In addition, they can also be used as novel protein-derived materials in a wide range of industries including cosmetics, raw materials of microcapsules and carriers of immobilized enzymes.
As enzymes having a possibility of increasing molecular weight of protein by cross-linking reaction, transglutaminase, lysyl oxidase, protein disulfide-isomerase, protein-disulfide reductase, sulfhydryl oxidase, lipoxygenase, polyphenol oxidase (tyrosinase) and peroxidase have been known (Matheis and Whitaker,
J. Food Biochemistry,
11, 309-327, 1987).
Among the above-described enzymes having a possibility of increasing molecular weight of protein by cross-linking reaction, a method for cross-linking protein by transglutaminase is well known. It is also known that this method has been broadly used mainly in the field of food processing based on the discovery of an inexpensive microbial transglutaminase which does not require the presence of calcium for the reaction (JP-B-6-65280 (the term “JP-B” as used herein means an “examined Japanese patent publication”),
Agric. Biol. Chem., vol.
69, no. 10, pp. 1301-1308).
The protein cross-linking reaction by transglutaminase, however, has the following problem. That is, since transglutaminase is an enzyme which forms an intramolecular or intermolecular bridge structure of protein as a result of the acyl rearrangement reaction generated between the &ggr;-carboxyl group of glutamine residue and the &egr;-amino group of lysine residue in a protein, some species of protein can hardly become the substrate for the enzyme due to insufficient glutamine residues or lysine residues. For example, albumin proteins cannot be used as the substrate for transglutaminase under their native state.
Thus, although a possibility of using several enzymes as the enzyme-aided protein cross-linking method has been indicated, most of them are not sufficiently useful in their supplying amounts, costs, easiness in the purification, and the like. Even if the microbial transglutaminase is used for the cross-linking method, its application is limited because of a problem that the reaction does not occur in some protein species. In consequence, great concern has been directed toward the development of a protein cross-linking method which uses other enzymes.
SUMMARY OF THE INVENTION
The inventor of the present invention has conducted extensive studies on the the possiblity of cross-linking reaction of protein using a multi-copper oxidase such as laccase, bilirubin oxidase, ascorbic acid oxidase and ceruloplasmin and, as a result, found that protein can be cross-linked by the use of these enzymes. Based on the finding of the novel method for cross-linking protein using a multi-copper oxidase such as laccase, bilirubin oxidase, ascorbic acid oxidase or ceruloplasmin, the present invention provides a protein cross-linking method which has a completely different reaction mechanism from that of transglutaminase and which can produce gelled protein having new physical properties and characteristics, with expanding the range of protein species to be treated which were limited in the known protein cross-linking method by transglutaminase.
The present inventors have made screening of a broad range of natural resources for finding a novel enzyme which can increase molecular weight of protein and makes it to gel by a cross-linking reaction and, as a result, found that certain enzymes classified as multi-copper oxidase can increase molecular weight of protein to make it gel by directly acting upon the protein, thus resulting in the accomplishment of the present invention.
REFERENCES:
patent: 5899212 (1999-05-01), Sorensen et al.
patent: 379 606 (1990-08-01), None
patent: 96/16165 (1996-05-01), None
Saari et al. J. Food Biochem. (1995), vol. 19, pp. 321-327.*
Faergemand M et al: “Cross-linking of whey proteins by enzymatic oxidation” Journal of Agricultural Food Chemistry, vol. 46, No. 4, Mar. 18, 1998, p. 1326-1333 XO000867797; *p. 1327, paragraph 1-3; *p. 1329, paragraph 4—p. 1330; paragraph 2: figures 5, 6; Table 1; *p. 1332, paragraph 2, 3.
Figueroa-Espinoza M C et al: “Oxidative Cross-Linking of Pentosans by a Fungal Laccase and Horseradish Peroxidase: Mechanism of Linkage between Feruloylated Arabinoxylans” Cereal Chemistry, US, American Association of Cereal Chemists. Minneapolis, vol. 75, No. 2, Mar. 1, 1998, pp. 259-265 XP000739677; ISSN: 0009-0352; *p. 259-p. 260; *p. 263; figure 2.
Matheis, G and Whitaker J.R.: “A review: Enzymatic cross-linking of proteins applicable to foods” Journal of Food Biochemistry, vol. 11, No. 4, 1987, p. 309-327 XOP000870468; Connecticut, USA; *p. 309-320.
Cooper et al., Biochem. Biophys. Res. Commun., 112(1); 161-167 (1983).
Amano Pharmaceutical Co. Ltd.
Coe Susan D.
Prats Francisco
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