Membrane Protein Stability

描述（由申请人提供）：膜蛋白在许多细胞过程中发挥重要作用。拟议研究的总体目标是了解蛋白质折叠过程的物理基础和膜蛋白结构的生物物理基础。目前，这两个都不是很好地了解膜蛋白。 热力学上，我们的工作解决了疏水性在膜蛋白折叠中发挥的关键作用。在前一个授予期间，我们开发了一种新的疏水性规模，测量侧链转移自由能从水膜中心使用一个真实的双层和一个真实的，折叠膜蛋白。基于这一成果，我们现在建议测试这个尺度的通用性：（1）通过使用不同的膜蛋白支架测量侧链转移自由能;（2）通过确定双层中的掩埋程度如何调节水到双层的转移自由能;以及（3）通过设计我们的蛋白质支架，用于测量pH值的函数，以解决可电离基团突变的能量后果如何随pH值的变化而变化。充电状态 在动力学上，我们发现在前一个赠款期间，E。大肠杆菌脂头 这些基团可以作为能量势，将膜蛋白从错误的（内）膜中分选出来，并朝向正确的（外）膜位置。我们建议在第四个目标，解剖的生物物理基础，这种排序通过确定的动力学寿命和构象和激活能折叠诱导的E。含大肠杆菌的脂质头部基团。

DESCRIPTION (provided by applicant): Membrane proteins play essential roles in many cellular processes. The overall goal of the proposed research is to understand the physical basis of the protein folding process and the biophysical basis of membrane protein structures. At present, neither of these is well understood for membrane proteins. Thermodynamically, our work addresses the critical role that hydrophobicity plays in membrane protein folds. In the previous granting period, we developed a novel hydrophobicity scale that measures side-chain transfer free energies from water to the membrane center using a real bilayer and a real, folded membrane protein. Based on this achievement, we now propose to test the generality of this scale (1) By measuring side-chain transfer free energies using distinct membrane protein scaffolds; (2) By determining how extent-of-burial in the bilayer modulates water to bilayer transfer free energies; and (3) By engineering of our protein scaffold for measurements as a function of pH to address how the energetic consequences of ionizable group mutations vary with charge state. Kinetically, we discovered in the previous grant period that E. coli lipid head groups may act as energetic potentials that sort membrane proteins away from the wrong (inner) membranes and towards the correct (outer) membrane locations. We propose in a fourth aim to dissect the biophysical basis for this sorting by determining the kinetic lifetimes and conformations and activation energies to folding induced by E. coli-containing lipid head groups.