Heat-induced redistribution of disulfide bonds in milk proteins. 2. Disulfide bonding patterns between bovine beta-lactoglobulin and kappa-casein.
Heat treatment of milk causes the heat-denaturable whey proteins to aggregate with kappa-casein (kappa-CN) via thiol-disulfide bond interchange reactions. The particular disulfide bonds that are important in the aggregates are uncertain, although Cys(121) of beta-lactoglobulin (beta-LG) has been implicated. The reaction at 60 degrees C between beta-LG A and an activated kappa-CN formed small disulfide-bonded aggregates. The tryptic peptides from this model system included a peptide with a disulfide bond between a Cys residue in the triple-Cys peptide [beta-LG(102-124)] and kappa-CN Cys(88) and others between kappa-CN Cys(88) or kappa-CN Cys(11) and beta-LG Cys(160). Only the latter two novel disulfide bonds were identified in heated (90 degrees C/20 min) milk. Application of computational search tools, notably MS2Assign and SearchXLinks, to the mass spectrometry (MS) and collision-induced dissociation (CID)-MS data was very valuable for identifying possible disulfide-bonded peptides. In two instances, peptides with measured masses of 4275.07 and 2312.07 were tentatively assigned to beta-LG(102-135):kappa-CN(11-13) and beta-LG A(61-69):kappa-CN(87-97), respectively. However, sequencing using the CID-MS data demonstrated that they were, in fact, beta-LG(1-40) and beta-LG(41-60), respectively. This study supports the notion that reversible intramolecular disulfide-bond interchange precedes the intermolecular interchange reactions.
Lowe
Edwin K
EK
Fonterra Research Centre, Private Bag 11-029, Palmerston North, New Zealand. edwin.lowe@fonterra.com
Anema
Skelte G
SG
Bienvenue
Annie
A
Boland
Michael J
MJ
Creamer
Lawrence K
LK
Jiménez-Flores
Rafael
R
D000595:Amino Acid Sequence; D000818:Animals; D002364:Caseins / Q000737:chemistry*; D002417:Cattle; D002851:Chromatography, High Pressure Liquid; D019281:Dimerization; D004220:Disulfides / Q000737:chemistry*; D004591:Electrophoresis, Polyacrylamide Gel; D006358:Hot Temperature; D007782:Lactoglobulins / Q000737:chemistry*; D013058:Mass Spectrometry; D008892:Milk / Q000737:chemistry; D008894:Milk Proteins / Q000737:chemistry*; D008969:Molecular Sequence Data; D010446:Peptide Fragments / Q000737:chemistry / Q000378:metabolism; D014357:Trypsin / Q000378:metabolism
D002364:Caseins; D004220:Disulfides; D007782:Lactoglobulins; D008894:Milk Proteins; D010446:Peptide Fragments; D014357:Trypsin