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Irreversible heat denaturation of bovine α-lactalbumin

Published online by Cambridge University Press:  01 June 2009

Lesley C. Chaplin  and
Richard L. J. Lyster
Lesley C. Chaplin
Affiliation:
AFRC Food Research Institute (University of Reading), Shinfield, Reading RG2 9AT, UK
Richard L. J. Lyster
Affiliation:
AFRC Food Research Institute (University of Reading), Shinfield, Reading RG2 9AT, UK
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Summary

The irreversible heat denaturation of α-lactalbumin (α-la) in 0·1 M-phosphate, pH 7·0, at 100 °C was studied using polyacrylamide-gel electrophoresis (PAGE). PAGE revealed two groups of bands, one moving faster than native α-la and one slower, in addition to some denatured protein which remained at the origin and some residual native α-la. The faster group had unchanged molecular weight, but an increase in charge, partly due to hydrolysis of glutamine and asparagine residues. The slower group was shown by two-dimensional sodium dodecyl sulphate-PAGE to be oligomers of denatured α-la; formation of the smaller oligomers preceded the larger ones. The oligomers reverted to monomers in the presence of dithiothreitol, showing that they were disulphide-linked aggregates of denatured α-la. Immuno-blots of the gels showed that both fast and slow groups of bands had irreversibly lost most of the antigenicity of the native protein.

Type
Original Articles
Information
Journal of Dairy Research , Volume 53 , Issue 2 , May 1986 , pp. 249 - 258
Copyright
Copyright © Proprietors of Journal of Dairy Research 1986

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References

REFERENCES

Arai, K. & Watanabe, S. 1968 A study of troponin, a myofibrillar protein from skeletal muscle. Journal of Biological Chemistry 243 5670–5678CrossRefGoogle ScholarPubMed
Aschaffenburg, R. & Drewry, J. 1957 Improved method for the preparation of crystalline β-lactoglobulin and α-lactalbumin from cow's milk. Biochemical Journal 65 273–277CrossRefGoogle ScholarPubMed
Baer, A., Oroz, M. & Blanc, B. 1976 Serological studies on heat-induced interactions of α-lactalbumin and milk proteins. Journal of Dairy Research 43 419–432CrossRefGoogle Scholar
Browne, W. J., North, A. C. T., Phillips, D. C., Brew, K., Vanaman, T. C. & Hill, R. L. 1969 A possible three-dimensional structure of bovine α-lactalbumin based on that of hen's egg-white lysozyme. Journal of Molecular Biology 42 65–86CrossRefGoogle ScholarPubMed
Bryant, D. T. W. & Andrews, P. 1984 High-affinity binding of Ca2+ to bovine α-lactalbumin in the absence and presence of EGTA. Biochemical Journal 220 617–620CrossRefGoogle ScholarPubMed
Hawkes, R., Niday, E. & Gordon, J. 1982 A dot-immunobinding assay for monoclonal and other antibodies. Analytical Biochemistry 119 142–147CrossRefGoogle ScholarPubMed
Hillier, R. M. 1976 The quantitative measurement of whey proteins using polyacrylamide-gel electrophoresis. Journal of Dairy Research 43 259–265CrossRefGoogle ScholarPubMed
Hillier, R. M., Lyster, R. L. J. & Cheeseman, G. C. 1979 Thermal denaturation of α-lactalbumin and β-Mactoglobulin in cheese whey: effect of total solids concentration and pH. Journal of Dairy Research 46 103–111CrossRefGoogle Scholar
Hiraoka, Y., Segawa, T., Kuwajima, K., Sugai, S. & Murai, N. 1980 α-Lactalbumin: a calcium metallo protein. Biochemical and Biophysical Research Communications 95 1098–1104CrossRefGoogle Scholar
Jenness, R. 1970 Protein composition of milk. In Milk Proteins: Chemistry and Molecular Biology Vol. 1, p. 25 (Ed. McKenzie, H. A.). New York: Academic Press.Google Scholar
Laemmli, U. K. 1970 Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature 227 680–685CrossRefGoogle ScholarPubMed
Levieux, D. 1980 Heat denaturation of whey proteins: comparative studies with physical and immunological methods. Annates de Recherches Vétérinaires 11 89–97Google ScholarPubMed
Lyster, R. L. J. 1970 The denaturation of α-lactalbumin and β-lactoglobulin in heated milk. Journal of Dairy Research 37 233–243CrossRefGoogle Scholar
Pfeil, W. 1981 Thermodynamics of α-lactalbumin unfolding. Biophysical Chemistry 13 181–186CrossRefGoogle ScholarPubMed
Privalov, P. L. 1979 Stability of proteins: small globular proteins. Advances in Protein Chemistry 33 167–241CrossRefGoogle ScholarPubMed
Reardon, J., Foreman, J. A. & Searcy, R. L. 1966 New reactants for the colorimetric determination of ammonia. Clinica Chimica Acta 14 403–405CrossRefGoogle ScholarPubMed
Rüegg, M., Moor, U., Lukesoh, A. & Blanc, B. 1977 a In Applications of Calorimetry in Life Sciences pp. 59–73 (Eds. Lamprecht, I. and Schaarschmidt, B.). Berlin: Walter de GruyterGoogle Scholar
Rüegg, M., Moor, U. & Blanc, B. 1977b A calorimetric study of the thermal denaturation of whey proteins in simulated milk ultrafiltrate. Journal of Dairy Research 44 509–520CrossRefGoogle Scholar
Saravis, C. A. 1984 Improved blocking of nonspecific antibody binding sites on nitrocellulose membranes. Electrophoresis 5 54–55CrossRefGoogle Scholar
Schnack, U. & Klostermeyer, H. 1980 Thermal decomposition of α-lactalbumin. 1. Destruction of cystine residues. Milchunssenschaft 35 206–208Google Scholar