Ropson Ira J, Boyer Joshua A, Dalessio Paula M
Department of Biochemistry and Molecular Biology, The Pennsylvania State University College of Medicine, Hershey, Pennsylvania 17033, USA.
Biochemistry. 2006 Feb 28;45(8):2608-17. doi: 10.1021/bi052091o.
Much of the recent effort in protein folding has focused on the possibility that residual structures in the unfolded state may provide an initiating site for protein folding. This hypothesis is difficult to test because of the weak stability and dynamic behavior of these structures. This problem has been simplified for intestinal fatty acid binding protein (IFABP) by incorporating fluorinated aromatic amino acids during synthesis in Escherichia coli. Only the labeled residues give signals by (19)F NMR, and the 1D spectra can be assigned in both the native and unfolded states by site-directed mutagenesis. One of the two tryptophans (W82), one of the four tyrosines (Y70), and at least four of the eight phenylalanines (including F68 and F93) of IFABP are involved in a structure that is significantly populated at concentrations of urea that unfold the native structure by fluorescence and CD criteria. These residues are nonlocal in sequence and also contact each other in the native structure. Thus, a template of nativelike hydrophobic contacts in the unfolded state may serve as an initiating site for folding this beta-sheet protein.
未折叠状态下的残余结构可能为蛋白质折叠提供起始位点。由于这些结构稳定性较弱且具有动态行为，这一假说难以验证。通过在大肠杆菌合成过程中引入氟化芳香族氨基酸，肠道脂肪酸结合蛋白（IFABP）的这一问题得到了简化。只有标记的残基能通过（19）F NMR给出信号，并且通过定点诱变可在天然状态和未折叠状态下对一维光谱进行归属。IFABP的两个色氨酸之一（W82）、四个酪氨酸之一（Y70）以及八个苯丙氨酸中的至少四个（包括F68和F93）参与了一种结构，在通过荧光和圆二色性标准使天然结构展开的尿素浓度下，该结构大量存在。这些残基在序列上不相邻，但在天然结构中相互接触。因此，未折叠状态下类似天然的疏水接触模板可能作为折叠这种β-折叠蛋白的起始位点。
