Hostname: page-component-586b7cd67f-tf8b9 Total loading time: 0 Render date: 2024-11-22T22:12:55.788Z Has data issue: false hasContentIssue false
  • English
  • Français

Proteolytic specificity of chymosin on bovine αs1,-casein

Published online by Cambridge University Press:  01 June 2009

Paul L. H. McSweeney ,
Norman F. Olson ,
Patrick F. Fox ,
Aine Healy  and
Peter Højrup
Paul L. H. McSweeney
Affiliation:
Center for Dairy Research, University of Wisconsin-Madison, Madison, WI 53706, USA
Norman F. Olson
Affiliation:
Center for Dairy Research, University of Wisconsin-Madison, Madison, WI 53706, USA
Patrick F. Fox
Affiliation:
Department of Food Chemistry, University College, Cork, Irish Republic National Food Biotechnology Centre, University College, Cork, Irish Republic
Aine Healy
Affiliation:
National Food Biotechnology Centre, University College, Cork, Irish Republic
Peter Højrup
Affiliation:
Department of Molecular Biology, University of Odense, Campusvej 55, DK-5230, Odense M., Denmark
Get access Rights & Permissions [Opens in a new window]

Summary

The proteolytic specificity of chymosin (EC 3.4.23.4) on bovine αs1-casein at 30°C in phosphate buffer, pH 6·5 and at pH 5·2 in the presence of 5% (w/v) NaCl was investigated. Peptides (pH 4·6-soluble) were isolated by reversed-phase HPLC and identified from their amino acid sequence; the identity of some peptides was confirmed by mass spectrometry and/or amino acid composition. The small peptides produced at pH 6·5 were Arg1–Phe23, Phe24–Phe28, Phe24–Leu40(?), Phe150–Phe153, Phe150–Leu156, Tyr154–Tyr159, Tyr154–Trp164, Asp157–Trp164 and Tyr165–Trp199. The same peptides, except Tyr154–Trp164, were produced at pH 5·2 in the presence of NaCl and, in addition, the peptides Arg1–Leu11, Phe24–Phe32, Lys102–Leu142, Ala143-Leu149 and Tyr165-Phe179. The rates of production of individual peptides differed under the two conditions studied but Arg1-Phe23 and Tyr165–Trp199 were the first and second peptides produced under both conditions. Pathways are proposed to interpret the proteolysis of αs1-casein in solution under the conditions of this study.

Type
Original Articles
Information
Journal of Dairy Research , Volume 60 , Issue 3 , August 1993 , pp. 401 - 412
Copyright
Copyright © Proprietors of Journal of Dairy Research 1993

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

Andrews, A. T. 1983 Proteinases in normal bovine milk and their action on caseins. Journal of Dairy Research 50 45–55CrossRefGoogle ScholarPubMed
Barbano, D. M. & Rasmussen, R. R. 1992 Cheese yield performance of fermentation-produced chymosin and other milk coagulants. Journal of Dairy Science 75 1–12CrossRefGoogle Scholar
Berridge, N. J. 1956 Crystalline versus crude rennet in cheesemaking Proceedings of the XIV International Dairy Congress, Rome, 2 59–66Google Scholar
Bines, V. E., Young, P. & Law, B. A. 1989 Comparison of Cheddar cheese made with a rccombinant calf chymosin and with standard calf rennet. Journal of Dairy Research 56 657–664CrossRefGoogle Scholar
Blakesley, R. W. & Boezi, J. A. 1977 A new staining technique for proteins in polyacrylamide gels using Coomassin brilliant blue G250. Analytical Biochemistry 82 580–582CrossRefGoogle ScholarPubMed
Carles, C. & Ribadeau Dumas, B. 1985 Kinetics of the action of chymosin (rennin) on a peptide bond of bovine αs1-casein. Comparison of the behaviour of this substrate with that of β- and κ0-caseins. FEBS Letters 185 282–286CrossRefGoogle Scholar
Creamer, L. K. 1974 Preparation of αs1-casein A. Journal of Dairy Science 57 341–344CrossRefGoogle Scholar
Creamer, L. K. & Olson, N. F. 1982 Rheological evaluation of maturing Cheddar cheese. Journal of Food Science 47 631–636, 646CrossRefGoogle Scholar
Creamer, L. K. & Richardson, B. C. 1974 Identification of the primary degradation product of αs1-casein in Cheddar cheese. New Zealand Journal of Dairy Science and Technology 9 9–13Google Scholar
Delfour, A., Jolles, J., Alais, C. & Jolles, P. 1965 Caseinoglycopeptides: characterization of a methionine residue and of the N-terminal sequence. Biochemical and Biophysical Research Communications 19 452–455CrossRefGoogle ScholarPubMed
Dunn, B. M., Valler, M. J., Rolph, C. E., Foundling, S. I., Jimenez, M. & Kay, J. 1987 The pH dependence of the hydrolysis of chromogenic substrates of the type, Lys–Pro–Xaa–Yaa–Phe– (N02)Phe–Arg–Leu, by selected aspartic proteinascs: evidence for specific interactions in subsites S3 and S2. Biochimica et Biophysica Acta 913 122–130CrossRefGoogle Scholar
Fish, J. C. 1957 Activity and specificity of rennin. Nature 180 345CrossRefGoogle ScholarPubMed
Fox, P. F. 1988 Rennets and their action in cheese manufacture and ripening. Biotechnology and Applied Biochemistry 10 522–535Google Scholar
Fox, P. F. 1989 Proteolysis during cheese manufacture and ripening. Journal of Dairy Science 72 1379–1400CrossRefGoogle Scholar
Green, M. L., Angal, S., Lowe, P. A. & Marston, F. A. O. 1985 Cheddar cheesemaking with recombinant calf chymosin synthesized in Eschcrichia coli. Journal of Dairy Research 52 281–286CrossRefGoogle Scholar
Grosclaude, F., Mahé, M.-F. & Ribadeau-Dumas, B. 1973 [Primary structure of bovine αs1- and β-casein. Correction.] European Journal of Biochemistry 40 323–324CrossRefGoogle Scholar
Hicks, C. L., O'Leary, J. & Bucy, J. 1988 Use of recombinant chymosin in the manufacture of Cheddar and Colby cheese. Journal of Dairy Science 71 1127–1131CrossRefGoogle Scholar
Hill, R. D. 1963 The preparation of κ-casein. Journal of Dairy Research 30 101–107CrossRefGoogle Scholar
Hill, R. D., Lahav, E. & Givol, D. 1974 A rennin-sensitive bond in αs1 B-casein. Journal of Dairy Research 41 147–153CrossRefGoogle ScholarPubMed
Le Bars, D. & Ghipon, J.-C. 1989 Specificity of plasmin towards bovine αs2-casein. Journal of Dairy Research 56 817–821CrossRefGoogle Scholar
McGann, T. C. A., Mathiassen, A. & O'Connell, J. A. 1972 Applications of the Pro-Milk Mk II. Part III. Rapid estimation of casein in milk and protein in whey. Laboratory Practice 21 628–631, 650Google Scholar
Meisel, H. 1987 [Characterization of rennet preparations produced by genetic engineering in comparison to calf rennet. 1. Materials and methods.] Milchwissenschaft 42 787–789Google Scholar
Meisel, H. 1988 [Characterization of rennet preparations produced by genetic engineering in comparison to calf rennet. 2. Results.] Milchwissenschaft 43 71–75Google Scholar
Mercier, J.-C., Grosclaude, F. & Ribadeau-Dumas, B. 1971 [Primary structure of bovine αs1-casein. Complete sequence.] European Journal of Biochemistry 23 41–51CrossRefGoogle Scholar
Mulvihill, D. M. & Fox, P. F. 1977 Proteolysis of αs1-casein by chymosin: influence of pH and urea. Journal of Dairy Research 44 533–540CrossRefGoogle Scholar
Mulvihill, D. M. & Fox, P. F. 1979 Proteolytic specificity of chymosin on bovine αs1-casein. Journal of Dairy Research 46 641–651CrossRefGoogle Scholar
Ng-Kwai-Hang, K. F., Monardes, H. G. & Hayes, J. F. 1990 Association between genetic polymorphism of milk proteins and production traits during three lactations. Journal of Dairy Science 73 3414–3420CrossRefGoogle Scholar
Nielsen, P. F., Klarskov, K., Højrup, P. & Roepstorff, P. 1988 Optimization of sample preparation for plasma desorption mass spectrometry of peptides and proteins using a nitrocellulose matrix. Biomedical and Environmental Mass Spectrometry 17 355–362CrossRefGoogle Scholar
O'Keeffe, A. M., Fox, P. F. & Daly, C. 1978 Proteolysis in Cheddar cheese: role of coagulant and starter bacteria. Journal of Dairy Research 45 465–477CrossRefGoogle Scholar
O'Keeffe, R. B., Fox, P. F. & Daly, C. 1976 Contribution of rennet and starter proteases to proteolysis in Cheddar cheese. Journal of Dairy Research 43 97–107CrossRefGoogle Scholar
Olson, N. F. 1990 The impact of lactic acid bacteria on cheese flavor. FEMS Microbiology Reviews 87 131–147CrossRefGoogle Scholar
O'Sullivan, M. & Fox, P. F. 1991 Evaluation of microbial chymosin from genetically engineered Kluveromyces lactis. Food Biotechnology 5 19–32CrossRefGoogle Scholar
Pahkala, E., Plhlanto-Leppälä, A., Laukkanen, M. & Antila, V. 1989 Decomposition of milk proteins during the ripening of cheese. 1. Enzymatic hydrolysis of αs-casein. Meijeritieteellinen Aikakauskirja 47 39–47Google Scholar
Pélissier, J.-P., Mercier, J.-C. & Ribadeau Dumas, B. 1974 [Study of the proteolysis of bovine αs1 and β-caseins by rennet. Specificity of action. Bitter peptides released.] Annales de Biologie Animale, Biochimie Biophysique 14 343–362Google Scholar
Rothe, G. A. L., Harboe, M. K. & Martiny, S. C. 1977 Quantification of milk-clotting enzymes in 40 commercial bovine rennets, comparing rocket immunoeleotrophoresis with an activity ratio assay. Journal of Dairy Research 44 73–77CrossRefGoogle Scholar
Van Den Berg, G. & De Koning, P. J. 1990 Gouda cheesemaking with purified calf chymosin and microbially produced chymosin. Netherlands Milk and Dairy Journal 44 189–205Google Scholar
Van Eenennaam, A. & Medrano, J. F. 1991 Milk protein polymorphisms in California dairy cattle. Journal of Dairy Science 74 1730–1742CrossRefGoogle Scholar
Visser, S. & Slangen, K. J. 1977 On the specificity of chymosin (rennin) in its action on bovine β-casein. Netherlands Milk and Dairy Journal 31 16–30Google Scholar