Ragona L, Confalonieri L, Zetta L, De Kruif K G, Mammi S, Peggion E, Longhi R, Molinari H
Laboratorio NMR, ICM, CNR, Milano, Italy.
Biopolymers. 1999 May;49(6):441-50. doi: 10.1002/(SICI)1097-0282(199905)49:6<441::AID-BIP2>3.0.CO;2-A.
Bovine beta-lactoglobulin represents an interesting example of context-dependent secondary structure induction. In fact, secondary structure predictions indicated that this beta-barrel protein has a surprisingly high alpha-helical preference, which was retained for short fragments. Cooperative transitions from the native beta-sheet to alpha-helical structures were additionally induced by organic solvents, in particular trifluoroethanol. As a result of this high alpha-helical preference, it has been proposed that non-native alpha-helical intermediates could be formed in the unfolding pathway of this protein. In order to provide a better understanding of the processes that underlie conformational plasticity in this protein, CD measurements in the presence of increasing amounts of urea and in the presence of organic solvents were performed. Urea unfolding studies, performed at pH 2.1 and 37 degrees C, revealed an apparent two-state transition, and afforded no evidence of non native alpha-helical intermediates. The protein treated with up to 6M urea, refolded to the native structure, while treatment with higher molar concentration urea, lead to partial misfolding. A 29-mer peptide covering the region of strands a and b of the intact protein, characterized by the presence of 4/3 heptad repeats, was synthesized and studied by CD in the presence of different solvents. On the basis of the obtained results, a mechanism was proposed to explain the structural transition from the beta to alpha structure, provoked by organic solvents in the intact protein.
牛β-乳球蛋白是一个依赖于环境的二级结构诱导的有趣例子。事实上，二级结构预测表明，这种β-桶状蛋白具有出人意料的高α-螺旋偏好，这种偏好对于短片段来说是保留的。有机溶剂，特别是三氟乙醇，还会诱导从天然β-折叠到α-螺旋结构的协同转变。由于这种高α-螺旋偏好，有人提出在这种蛋白质的解折叠途径中可能形成非天然的α-螺旋中间体。为了更好地理解这种蛋白质构象可塑性的潜在过程，我们在存在越来越多尿素的情况下以及在有机溶剂存在的情况下进行了圆二色性测量。在pH 2.1和37℃下进行的尿素解折叠研究揭示了明显的两态转变，并且没有提供非天然α-螺旋中间体的证据。用高达6M尿素处理的蛋白质重新折叠成天然结构，而用更高摩尔浓度的尿素处理则导致部分错误折叠。合成了一个覆盖完整蛋白质的a链和b链区域的29肽，其特征是存在4/3个七肽重复序列，并在不同溶剂存在的情况下通过圆二色性进行了研究。根据获得的结果，提出了一种机制来解释完整蛋白质中有机溶剂引发的从β结构到α结构的结构转变。
