Stanley Ann Marie, Fleming Karen G
T.C. Jenkins Department of Biophysics, Johns Hopkins University, 3400 North Charles Street, Baltimore, MD 21218, USA.
J Mol Biol. 2005 Apr 8;347(4):759-72. doi: 10.1016/j.jmb.2005.01.059.
The epidermal growth factor receptors (erbB) constitute an important class of single pass transmembrane receptors involved in the transduction of signals important for cell proliferation and differentiation. Receptor association is a key event in the signal transduction process, but the molecular basis of this interaction is not fully understood. Previous biochemical and genetic studies have suggested that the single transmembrane helices of these receptor proteins might play a role in stabilizing the receptor complexes. To determine if the erbB transmembrane domains could provide a driving force to stabilize the receptor dimers, we carried out a thermodynamic study of these domains expressed as C-terminal fusion proteins with staphylococcal nuclease. Similar fusion constructs have been used successfully to investigate the oligomerization and association thermodynamics of a number of transmembrane sequences, including that of glycophorin A. Using SDS-PAGE analysis and sedimentation equilibrium analytical ultracentrifugation, we do not find strong, specific homo or hetero-interactions between the transmembrane domains of the erbB receptors in micellar solutions. Our results indicate that any preferential interactions between these domains in micellar solutions are extremely modest, of the order of 1 kcal mol(-1) or less. We applied a thermodynamic formalism to assess the effect of weakly interacting TM segments on the behavior of a covalently attached soluble domain. In the case of the ligand-bound EGFR ectodomain, we find that restriction of the ectodomain to the micellar phase by a hydrophobic TM, even in the absence of strong specific interactions, is largely sufficient to account for the previously reported increase in dimerization affinity.
表皮生长因子受体（erbB）构成了一类重要的单次跨膜受体，参与对细胞增殖和分化至关重要的信号转导。受体缔合是信号转导过程中的关键事件，但这种相互作用的分子基础尚未完全了解。先前的生化和遗传学研究表明，这些受体蛋白的单个跨膜螺旋可能在稳定受体复合物中发挥作用。为了确定erbB跨膜结构域是否能提供稳定受体二聚体的驱动力，我们对以葡萄球菌核酸酶C末端融合蛋白形式表达的这些结构域进行了热力学研究。类似的融合构建体已成功用于研究许多跨膜序列的寡聚化和缔合热力学，包括血型糖蛋白A的跨膜序列。通过SDS-PAGE分析和沉降平衡分析超速离心，我们在胶束溶液中未发现erbB受体跨膜结构域之间有强烈的、特异性的同源或异源相互作用。我们的结果表明，这些结构域在胶束溶液中的任何优先相互作用都极其微弱，约为1千卡摩尔（-1）或更低。我们应用一种热力学形式来评估弱相互作用的跨膜片段对共价连接的可溶性结构域行为的影响。对于配体结合的表皮生长因子受体胞外结构域，我们发现，即使在没有强烈特异性相互作用的情况下，疏水跨膜结构域将胞外结构域限制在胶束相中，在很大程度上足以解释先前报道的二聚化亲和力增加的现象。
