Gratkowski H, Lear J D, DeGrado W F
The Johnson Research Foundation, Department of Biochemistry and Biophysics, School of Medicine, University of Pennsylvania, Philadelphia, PA 19104-6059, USA.
Proc Natl Acad Sci U S A. 2001 Jan 30;98(3):880-5. doi: 10.1073/pnas.98.3.880.
The forces stabilizing the three-dimensional structures of membrane proteins are currently not well understood. Previously, it was shown that a single Asn side chain in a transmembrane segment can mediate the dimerization and trimerization of a variety of hydrophobic helices. Here, we examine the tendencies of a representative set of amino acids (Asn, Gln, Asp, Glu, Lys, Ala, Val, Leu, Ser, Thr) to direct the oligomerization of a model transmembrane helix. The model peptide is entirely hydrophobic throughout a 20-residue segment and contains a single central site for the introduction of various amino acid "guests." Analytical ultracentrifugation and gel electrophoresis were used to determine the stoichiometry and free energy of association of the entire set of peptides within micelles. Variants with two polar atoms at the guest site-Asn, Gln, Asp, and Glu-formed stable trimers, whereas residues with one or fewer polar atoms showed a much weaker tendency to associate. The data are examined in light of the frequencies of occurrence of various amino acid side chains in membrane proteins and provide insight into the role of polar interactions in directing transmembrane helix association. These data also suggest an approach to the design of variants of natural single-span transmembrane proteins with various potentials to associate in the bilayer.
目前，稳定膜蛋白三维结构的作用力尚未得到充分理解。此前研究表明，跨膜片段中的单个天冬酰胺（Asn）侧链可介导多种疏水螺旋的二聚化和三聚化。在此，我们研究了一组具有代表性的氨基酸（天冬酰胺、谷氨酰胺、天冬氨酸、谷氨酸、赖氨酸、丙氨酸、缬氨酸、亮氨酸、丝氨酸、苏氨酸）引导模型跨膜螺旋寡聚化的倾向。该模型肽在20个残基的片段中完全疏水，并且包含一个用于引入各种氨基酸“客体”的单一中心位点。使用分析型超速离心和凝胶电泳来确定胶束内整套肽的化学计量和缔合自由能。在客体位点带有两个极性原子的变体——天冬酰胺、谷氨酰胺、天冬氨酸和谷氨酸——形成了稳定的三聚体，而带有一个或更少极性原子的残基显示出弱得多的缔合倾向。根据膜蛋白中各种氨基酸侧链的出现频率对数据进行了研究，从而深入了解极性相互作用在引导跨膜螺旋缔合中的作用。这些数据还提出了一种设计天然单跨膜蛋白变体的方法，这些变体在双层中具有不同的缔合潜力。
