Plowman J E, Creamer L K, Liddell M J, Cross J J
Food Service Section, New Zealand Dairy Research Institute, Palmerston North, New Zealand.
J Dairy Res. 1997 Aug;64(3):377-97. doi: 10.1017/s0022029997002239.
The peptide Pro130-Thr-Ser-Thr-Pro-Thr-Ile-Glu-Ala-Val-Glu140- Ser-Thr-Val-Ala-Thr-Leu-GLu-Ala-Ser-Pro150-Glu-Val-Ile, which corresponds to residues 130-150 of kappa-casein B, was synthesized and the conformation of the peptide in solution investigated by circular dichroism (CD) spectroscopy, structure prediction algorithms and 1H-nuclear magnetic resonance spectroscopy. In a solution containing the structure-enhancing solvent trifluoroethanol the CD spectrum was typical of a peptide in the alpha-helical conformation and nuclear magnetic resonance showed that the amino acids between Ile136 and Ser149 (kappa-casein numbering) were predominantly in the alpha-helical conformation but that Pro130 to Thr135 and Pro150 to Ile153 were not. In addition, Thr133-Pro134 and Ser-149-Pro150 were primarily in the trans conformation, the residues from Thr131 to Thr135 were in unordered structures and the residues from Glu151 to Ile153 were in an extended conformation. Residues Glu137 to Glu140 and Thr145 to Ala148 also displayed some 3(10)-helix character. When the peptide was dissolved in 10 mM-cetyltrimethylammonium chloride solution at pH 6, the CD spectra indicated that the proportion of helical structure was comparable to that of the peptide in trifluoroethanol solution (400 ml/l), whereas when the peptide was dissolved in buffer alone in 10 mM-SDS solution, the CD spectra were consistent with a low helical content. Acidification of these solutions to pH 2.85 resulted in a slight increase in the helical content of the peptide in buffer and more markedly in buffer containing SDS. When the peptide was in 5 mM-CaCl2 solution at neutral pH, the CD spectrum indicated that some ordered structure was present. Taken together these results indicate that the ionizable residues Glu137, Glu140, Glu147 and Glu151 could be important in determining the stability of the putative helix. The structure predictions found that the sequence from Glu137 to Pro150 would be more likely to be in a helical than any other conformation in the intact bovine protein, but that pig, sheep and goat kappa-caseins did not give a prediction of a strongly helical region in this part of the molecule.
合成了对应于κ-酪蛋白B第130 - 150位残基的肽段Pro130 - Thr - Ser - Thr - Pro - Thr - Ile - Glu - Ala - Val - Glu140 - Ser - Thr - Val - Ala - Thr - Leu - Glu - Ala - Ser - Pro150 - Glu - Val - Ile，并通过圆二色性（CD）光谱、结构预测算法和1H - 核磁共振光谱研究了该肽段在溶液中的构象。在含有结构增强溶剂三氟乙醇的溶液中，CD光谱显示该肽段具有典型α - 螺旋构象，核磁共振表明Ile136和Ser149之间的氨基酸（κ-酪蛋白编号）主要处于α - 螺旋构象，但Pro130至Thr135和Pro150至Ile153并非如此。此外，Thr133 - Pro134和Ser - 149 - Pro150主要处于反式构象，Thr131至Thr135的残基处于无序结构，Glu151至Ile153的残基处于伸展构象。Glu137至Glu140和Thr145至Ala148的残基也显示出一些3(10)-螺旋特征。当该肽段溶解于pH 6的10 mM十六烷基三甲基氯化铵溶液中时，CD光谱表明螺旋结构的比例与该肽段在三氟乙醇溶液（400 ml/l）中的比例相当，而当该肽段仅溶解于10 mM SDS溶液的缓冲液中时，CD光谱显示螺旋含量较低。将这些溶液酸化至pH 2.85导致该肽段在缓冲液中的螺旋含量略有增加，在含有SDS的缓冲液中增加更为明显。当该肽段处于中性pH的5 mM CaCl2溶液中时，CD光谱表明存在一些有序结构。综合这些结果表明，可电离残基Glu137、Glu140、Glu147和Glu151在确定假定螺旋的稳定性方面可能很重要。结构预测发现，在完整的牛蛋白中，Glu137至Pro150的序列比其他任何构象更有可能处于螺旋构象，但猪、绵羊和山羊的κ-酪蛋白在该分子的这一部分并未预测到有强螺旋区域。
