Protease-induced aggregation of bovine alpha-lactalbumin: identification of the primary associating fragment.
Details in the hydrolysis of alpha-lactalbumin known to result in formation of highly ordered nanotubules, was investigated by incubation of solutions with 10 g alpha-lactalbumin/l with a specific protease from Bacillus licheniformis (BLP). After 50 min of incubation, soluble aggregates were formed, the concentration of which increased until precipitation occurred after 200 min. The latter aggregates were dissolved in urea or at low pH, like the nanotubules characteristic of gels formed by the action of BLP on alpha-lactalbumin at 100 g/l. On the molecular level, alpha-lactalbumin was initially cleaved into two large hydrophobic fragments with masses of 11.6 and 11.3 kDa, which in turn were cleaved in a stepwise manner into the ultimate fragment of 8.8 kDa. This fragment was the predominating component in the insoluble aggregates, and was identified as the sequences 26-37 and 50-113 of alpha-lactalbumin linked together by a disulphide bond. Cleavage of alpha-lactalbumin into this fragment probably created new hydrophobic surfaces and new calcium binding sites allowing its association into ordered structures.
Otte
Jeanette
J
Department of Dairy and Food Science, The Royal Veterinary and Agricultural University, Rolighedsvej 30, DK-1958 Frederiksberg C, Denmark. jo@kvl.dk
Ipsen
Richard
R
Ladefoged
Anne Marie
AM
Sørensen
John
J
D000595:Amino Acid Sequence; D000818:Animals; D001407:Bacillus / Q000201:enzymology; D002417:Cattle; D055598:Chemical Phenomena; D011232:Chemical Precipitation; D002627:Chemistry, Physical; D002851:Chromatography, High Pressure Liquid; D010450:Endopeptidases / Q000378:metabolism / Q000494:pharmacology*; D006868:Hydrolysis; D007768:Lactalbumin / Q000737:chemistry* / Q000378:metabolism; D008969:Molecular Sequence Data; D009391:Nephelometry and Turbidimetry; D010446:Peptide Fragments / Q000737:chemistry / Q000378:metabolism; D012995:Solubility; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D014508:Urea
D010446:Peptide Fragments; D014508:Urea; D007768:Lactalbumin; D010450:Endopeptidases