Characterization of multiply phosphorylated peptides selectively precipitated from a pancreatic casein digest.
Anticariogenic phosphopeptides, released during the hydrolysis of casein with trypsin, contain the cluster sequence Ser(P)-Ser(P)-Ser(P)-Glu-Glu and have commercial potential as toothpaste, mouthwash, and food additives for the prevention of dental caries. To develop a commercial-scale process for the production of these peptides, we have comprehensively characterized casein phosphopeptides that were selectively precipitated using Ca2+ and ethanol from an acid-clarified (pH 4.6) pancreatic casein hydrolysate. Casein was hydrolyzed using pancreatin at 50 degrees C for 2 h. The precipitate contained a series of casein phosphopeptides that were slightly truncated relative to tryptic casein phosphopeptides. The major casein phosphopeptides released by pancreatin were beta-CN-4P(f7-24), alpha s1-CN-5P(f61-78), and alpha s1-CN-5P(f59-78), all containing the cluster sequence. The truncation of the tryptic peptides beta-CN-4P(1-25) and alpha s1-CN-5P(f59-79) resulted from the chy-motryptic and carboxypeptidase activities of the pancreatin. The peptides containing the cluster sequence constituted 77.8 +/- 6.7 mol/100 mol of the total peptides that were selectively precipitated. This composition was not significantly different from that of casein phosphopeptides produced under identical conditions using trypsin. In conclusion, pancreatin should be a suitable enzyme preparation for the production of anticariogenic casein phosphopeptides on a commercial scale.
Adamson
N J
NJ
Biochemistry and Molecular Biology Unit, School of Dental Science, University of Melbourne, Australia.
Reynolds
E C
EC
D000595:Amino Acid Sequence; D002122:Calcium Chloride; D002364:Caseins / Q000737:chemistry* / Q000378:metabolism; D002851:Chromatography, High Pressure Liquid; D000431:Ethanol; D006863:Hydrogen-Ion Concentration; D008969:Molecular Sequence Data; D010179:Pancreas / Q000201:enzymology*; D010194:Pancreatin / Q000378:metabolism*; D010750:Phosphoproteins / Q000737:chemistry* / Q000378:metabolism; D013379:Substrate Specificity
D002364:Caseins; D010750:Phosphoproteins; D000431:Ethanol; D010194:Pancreatin; D002122:Calcium Chloride