Use of mass spectrometry To rapidly characterize the heterogeneity of bovine alpha-lactalbumin.
From a bovine whey protein fraction the nonglycosylated and glycosylated alpha-lactalbumin fractions were isolated by gel-permeation chromatography followed by reversed-phase high-performance liquid chromatography. Both fractions were studied by electrospray-ionization mass spectrometry (ESI-MS). For the nonglycosylated fraction, a mass of 14 178 Da was found, which was in accordance with the known amino acid sequence of the protein. The glycosylated fraction appeared to be a mixture of components with mass values ranging from ca. 15 840 to 16 690 Da. Given the published carbohydrate composition of the whole glyco-alpha-lactalbumin fraction, these masses could be matched to those of 14 differently glycosylated forms of alpha-lactalbumin. Further support for these forms was obtained from the results of a separate mass spectrometric analysis of the mixture of oligosaccharides released from the protein by treatment with peptide-N(4)-(N-acetyl-beta-glucosaminyl)asparagine amidase F. ESI-MS was found to be a powerful tool to gain insight into the composition of the complex mixture of glycoforms of alpha-lactalbumin without the need of further purification of these forms or of the oligosaccharides released thereof.
Slangen
C J
CJ
Department of Product Technology, NIZO Food Research, P.O. Box 20, 6710 BA Ede, The Netherlands.
Visser
S
S
D000818:Animals; D002236:Carbohydrate Conformation; D002417:Cattle; D002851:Chromatography, High Pressure Liquid; D006031:Glycosylation; D007768:Lactalbumin / Q000737:chemistry*; D013058:Mass Spectrometry
D007768:Lactalbumin