Jemth Per, Day Ryan, Gianni Stefano, Khan Faaizah, Allen Mark, Daggett Valerie, Fersht Alan R
MRC Centre for Protein Engineering, Hills Road, CB2 2QH Cambridge, UK.
J Mol Biol. 2005 Jul 8;350(2):363-78. doi: 10.1016/j.jmb.2005.04.067.
We have analysed the transition state of folding of the four-helix FF domain from HYPA/FBP11 by high-resolution experiment and simulation as part of a continuing effort to understand the principles of folding and the refinement of predictive methods. The major transition state for folding was subjected to a Phi-value analysis utilising 50 mutants. The transition state contained a nucleus for folding centred around the end of helix 1 (H1) and the beginning of helix 2 (H2). Secondary structure in this region was fully formed (PhiF=0.9-1) and tertiary interactions were well developed. Interactions in the distal part of the native structure were weak (PhiF=0-0.2). The hydrophobic core and other parts of the protein displayed intermediate Phi-values, suggesting that interactions coalesce as the end of H1 and beginning of H2 are in the process of being formed. The distribution of Phi-values resembled that of barnase, which folds via an intermediate, rather than that of CI2 which folds by a concerted nucleation-condensation mechanism. The overall picture of the transition state structure identified in molecular dynamics simulations is in qualitative agreement, with the turn connecting H1 and H2 being formed, a loosened core, and H4 partially unfolded and detached from the core. There are some differences in the details and interpretation of specific Phi-values.
作为理解折叠原理和完善预测方法的持续努力的一部分，我们通过高分辨率实验和模拟分析了来自HYPA/FBP11的四螺旋FF结构域的折叠过渡态。利用50个突变体对折叠的主要过渡态进行了Phi值分析。过渡态包含一个以螺旋1（H1）末端和螺旋2（H2）起始端为中心的折叠核心。该区域的二级结构已完全形成（PhiF = 0.9 - 1），三级相互作用也已充分发展。天然结构远端部分的相互作用较弱（PhiF = 0 - 0.2）。蛋白质的疏水核心和其他部分显示出中间Phi值，这表明随着H1末端和H2起始端的形成过程，相互作用逐渐聚集。Phi值的分布类似于通过中间体折叠的巴那斯酶，而不是通过协同成核凝聚机制折叠的CI2。分子动力学模拟中确定的过渡态结构的整体情况在定性上是一致的，连接H1和H2的转角正在形成，核心松散，H4部分展开并与核心分离。在特定Phi值的细节和解释上存在一些差异。
