Hostname: page-component-848d4c4894-x5gtn Total loading time: 0 Render date: 2024-06-02T09:18:56.163Z Has data issue: false hasContentIssue false

Immunological identification of milk-clotting enzymes

Published online by Cambridge University Press:  01 June 2009

Jean-Claude Collin
Affiliation:
Institut National de la Recherche Agronomique, CNRZ
Graciela Muset de Retta
Affiliation:
Institut National de la Recherche Agronomique, CNRZ
Patrice Martin
Affiliation:
Institut National de la Recherche Agronomique, CNRZ

Summary

The 6 most widely used milk-clotting enzymes, i.e. chymosin, bovine and porcine pepsins and proteinases from Mucor miehei, M. pusillus and Endothia parasitica, have been purified and used to prepare rabbit antisera against each of them. The antisera were adsorbed with appropriate cross-linked antigens to remove non-specific antibodies. Monospecific antisera thus obtained were used to identify enzymes eventually added to calf or bovine rennets, using the double radial immunodiffusion technique. The threshold of sensitivity was c. 1%, expressed as clotting activity. This qualitative method complements the quantitative procedure recently proposed for determining chymosin and bovine pepsin A in commercial extracts of bovine veils.

Type
Original Articles
Copyright
Copyright © Proprietors of Journal of Dairy Research 1982

Access options

Get access to the full version of this content by using one of the access options below. (Log in options will check for institutional or personal access. Content may require purchase if you do not have access.)

References

REFERENCES

Anson, M. L. 1938 The estimation of pepsin, trypsin, papain, and cathepsin with hemoglobin. Journal of General Physiology 22, 7989.CrossRefGoogle ScholarPubMed
Avrameas, S. & Ternynck, T. 1969 The cross-linking of proteins with glutaraldehyde and its use for the preparation of immunoadsorbents. Immunochemistry 6 5366.CrossRefGoogle ScholarPubMed
Cantagalli, P., Sordi, S., Piergiovanni, L. & Resmini, P. 1976 [Detection of calf rennet in kid and lamb rennets by immunochemical methods.] Scienza e Tecnica Lattiero-Casearia 27 2228Google Scholar
Collin, J. C., Martin, P., Garnot, P., Ribadeau Dumas, B. & Mocquot, G. 1981 Determination of chymosin and bovine pepsin A in commercial bovine rennets and pepsins. Milchwissenschaft 36 3235Google Scholar
De Koning, P. J. 1972 Identification of rennet substitutes. International Dairy Federation, Annual Bulletin part 4 115Google Scholar
De Koning, P. J. 1978 Method for the identification of rennet and rennet substitutes. International Dairy Federation, Annual Bulletin Document no. 108 5863Google Scholar
De Koning, P. J. & Draaisma, J. Th. M. 1973 Identification of different types of rennet by means of isoelectric focusing. Netherlands Milk and Dairy Journal 27 368378Google Scholar
Douillard, R. & Ribadeau Dumas, B. 1970 Détermination avec la caséine κ de l'activité protéolytique de la présure, de la pepsine de porc et des pepsines bovines. Bulletin de la Société de Chimie Biologique 52 14291445Google Scholar
Etoh, Y., Shoun, H., Beppu, T. & Arima, K. 1979 Physicochemical and immunochemical studies on similarities of acid proteases Mucor pusillus rennin and Mucor miehei rennin. Agricultural and Biological Chemistry 43 209215Google Scholar
Foltmann, B. & Pedersen, V. B. 1977 Comparison of the primary structures of acidic proteinases and of their zymogens. In Acid Proteases: Structure, Function, and Biology (Ed. Tang, J.) pp. 322New York, NY: Plenum PressCrossRefGoogle Scholar
Garnot, P., Thapon, J. L., Mathieu, C. M., Maubois, J. L. & Ribadeau Dumas, B. 1972 Determination of rennin and bovine pepsins in commercial rennets and abomasal juices. Journal of Dairy Science 55 16411650CrossRefGoogle Scholar
Green, M. L. 1977 Review of the progress of Dairy Science: milk coagulants. Journal of Dairy Research 44 159188CrossRefGoogle Scholar
Hagemayer, K., Fawwal, I. & Whitaker, J. R. 1968 Purification of protease from the fungus Endothia parasitica. Journal of Dairy Science 51 19161922CrossRefGoogle Scholar
Iwasaki, S., Yasui, T., Tamura, G. & Arima, K. 1967 Milk clotting enzymes from microorganisms. IV. Immunological studies on the enzyme properties Agricultural and Biological Chemistry 31 14271433CrossRefGoogle Scholar
Martin, P., Collin, J.-C., Garnot, P., Ribadeau Dumas, B. & Mocquot, G. 1981 Evaluation of bovine rennets in terms of absolute concentrations of chymosin and pepsin A. Journal of Dairy Research 48 447456CrossRefGoogle Scholar
Martin, P., Raymond, M.-N., Bricas, E., & Ribadeau Dumas, B. 1980 Kinetic studies on the action of Mucor pusillus, Mucor miehei acid proteases and chymosins A and B on a synthetic chromophoric hexapeptide. Biochimica et Biophysica Acta 612 410420CrossRefGoogle Scholar
Martiny, S. M. & Harboe, M. K. 1978 A system of analysis of any type of milk coagulant. 20th International Dairy Congress, Paris, E 445446Google Scholar
Molinari, G. 1979 [Identification of milk–clotting enzymes by isoelectric focusing.] Scienza e Tecnica Lattiero-Casearia 30 117121Google Scholar
Mulvihill, D. M. & Fox, P. F. 1977 Selective denaturation of milk coagulants in 5 M-urea. Journal of Dairy Research 44 319324CrossRefGoogle Scholar
Murakami, K. & Kobayashi, H. 1978 Rapid and large scale isolation of milk clotting enzymes by affinity chromatography. In Vth International Congress of Food Science and Technology Abstracts 230Google Scholar
O'Leary, P. A. & Fox, , 1974 A method for the quantitative analysis of the enzyme complement of commercial rennets. Journal of Dairy Research 41 381387CrossRefGoogle ScholarPubMed
Ottesen, M. & Rickert, W. 1970 The isolation and partial characterization of an acid protease produced by Mucor miehei. Comptes rendus des Travaux du Laboratoire Carlsberg 37 301325Google ScholarPubMed
Ouchterlony, Ö. 1948 In vitro method for testing the toxin producing capacity of diphtheria bacteria. Acta Pathologica et Microbiologica Scandinavica 25 186191CrossRefGoogle ScholarPubMed
Piazzi, S. E., Sordi, S., Resmini, P. & Cubadda, R. 1975 [Detection of the microbial rennets in calf rennets by immunoohemical method]. Scienza e Tecnica Lattiero-Casearia 26 2736Google Scholar
Prager, M. J. 1977 Differentiation of rennet from other milk-clotting enzymes by polyacrylamide gel electrophoresis. Journal of the Association of Official Analytical Chemists 60 13721374Google ScholarPubMed
Righetti, P. G., Molinari, B. M. & Molinari, G. 1977 Isoelectric focusing of milk-clotting enzymes. Journal of Dairy Research 44 6972CrossRefGoogle Scholar
Rothe, G. A. L., Axelsen, N. H., Jøhnk, P. & Foltmann, B. 1976 Immunochemical, chromatographic, and milk-clotting activity measurements for quantification of milk-clotting enzymes in bovine rennets. Journal of Dairy Research 43 8595CrossRefGoogle ScholarPubMed
Scheidegger, J. J. 1955 A micromethod of immuno-electrophoresis. International Archives of Allergy and Immunology 7 103110CrossRefGoogle ScholarPubMed
Shovers, J., Kornowski, R. & Fossum, G. 1973 Differential assay for milk coagulants in mixtures with porcine pepsin. Journal of Dairy Science 56 994997CrossRefGoogle Scholar
Wake, R. G. & Baldwin, R. L. 1961 Analysis of casein fractions by zone electrophoresis in concentrated urea. Biochimica et Biophysica Acta 47 225239CrossRefGoogle ScholarPubMed