Use of reducing/nonreducing two-dimensional electrophoresis for the study of disulfide-mediated interactions between proteins in raw and heated bovine milk.
The composition and interactions of proteins in bovine milk, and modifications resulting from milk storage and processing, are complex and incompletely understood. Analysis of the milk proteome can elucidate milk protein expression, structure, interaction, and modifications. Raw milk was analyzed by two-dimensional electrophoresis (isolelectric focusing followed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis) under reducing and nonreducing, or combined, conditions, followed by mass spectrometry of separated protein spots; a small number of high-abundance proteins, that is, caseins (alpha(S1)-, alpha(S2)-, beta-, kappa-, and gamma-), beta-lactoglobulin, alpha-lactalbumin, and serum albumin, represented the vast majority of protein spots on the two-dimensional electrophoretograms of raw milk samples, but some cross-linked protein complexes (mainly homopolymers of kappa-casein and alpha(S2)-casein but also some heteropolymeric complexes) were resolved under native/unheated conditions. When skim milk was heated to 90 degrees C for up to 10 min, the level of native whey proteins decreased in parallel with an increase in disulfide-linked complexes, including very complex heteropolymers, for example, casein/whey protein polymers containing multiple species. The analysis strategy used in this study reveals numerous disulfide-mediated interactions and can be proposed to analyze reduction/oxidation of milk and dairy product proteins following processing treatments applied for processing and storage.
Chevalier
François
F
Department of Food and Nutritional Sciences, University College Cork, Cork, Ireland. francois.chevalier@cea.fr
Hirtz
Christophe
C
Sommerer
Nicolas
N
Kelly
Alan L
AL
D000818:Animals; D002364:Caseins / Q000737:chemistry; D002417:Cattle; D004220:Disulfides / Q000737:chemistry*; D015180:Electrophoresis, Gel, Two-Dimensional; D006358:Hot Temperature; D008892:Milk / Q000737:chemistry*; D008894:Milk Proteins / Q000032:analysis / Q000737:chemistry*; D010084:Oxidation-Reduction; D000067816:Whey Proteins
D002364:Caseins; D004220:Disulfides; D008894:Milk Proteins; D000067816:Whey Proteins