Livney Yoav D, Verespej Edita, Dalgleish Douglas G
Department of Food Science, University of Guelph, Guelph, Ontario N1G 2W1, Canada.
J Agric Food Chem. 2003 Dec 31;51(27):8098-106. doi: 10.1021/jf034582q.
Intermolecular disulfide bond formation in pure beta-lactoglobulin (beta-Lg) B and in its 1:1 mixture with alpha-lactalbumin (alpha-La), heated at 85 degrees C for 10 min in solutions of low and high (100 mM NaCl) ionic strength and pH 6.0, was studied by reverse-phase HPLC and MALDI-TOF mass spectrometry. Disulfide bonding between beta-Lg monomers was more extensive than reported in the literature for a temperature of 68.5 degrees C, including formation of trimers connected by two of the three adjacent cysteines, C106/C119/C121. The participation of the different thiol groups in disulfide bonds appeared to depend on their location in the native structure, with surface-located cysteines more involved than internally located ones. This also applied to alpha-La-beta-Lg interactions, where the predominant participants were the surface-located alphaC111, alphaC120, alphaC61, and alphaC6. The least active participant was alphaC28, suggesting that it becomes sterically inaccessible during unfolding of the protein. High ionic strength apparently promoted disulfide bonding. The order of cysteine participation at the high ionic strength was similar to that at low ionic strength, with fewer native-location bonds observed and a lower activity of some groups, such as beta-C106/C119/C121 and alphaC61.
通过反相高效液相色谱法和基质辅助激光解吸电离飞行时间质谱法，研究了在低离子强度和高离子强度（100 mM NaCl）、pH 6.0的溶液中，于85℃加热10分钟的纯β-乳球蛋白（β-Lg）B及其与α-乳白蛋白（α-La）的1:1混合物中的分子间二硫键形成情况。β-Lg单体之间的二硫键形成比文献报道的68.5℃时更为广泛，包括由三个相邻半胱氨酸中的两个（C106/C119/C121）连接形成的三聚体。不同巯基参与二硫键形成似乎取决于它们在天然结构中的位置，表面定位的半胱氨酸比内部定位的半胱氨酸参与程度更高。这也适用于α-La-β-Lg相互作用，其中主要参与者是表面定位的αC111、αC120、αC61和αC6。活性最低的参与者是αC28，表明它在蛋白质解折叠过程中在空间上变得难以接近。高离子强度明显促进了二硫键的形成。高离子强度下半胱氨酸参与的顺序与低离子强度时相似，观察到的天然位置键较少，且一些基团（如β-C106/C119/C121和αC61）的活性较低。
