Flinders K T, Yu Y B, Flynn P F
Department of Pharmaceutics and Pharmaceutical Chemistry, University of Utah, Salt Lake City, UT 84112, USA.
J Pept Res. 2004 Jan;63(1):17-22. doi: 10.1046/j.1399-3011.2004.00099.x.
To assess the relative importance of backbone hydrogen bonding (H-bonding) vs. side chain hydrophobicity in protein structural formation, a method called side chain-backbone swap is proposed. Such a method swaps the side chain and backbone portions of certain amino acid residues, such as Asp, Glu, Asn, Gln, Lys, and Arg. Such a swap retains the sequence of a polypeptide and preserves the identity of the backbone linkage. On the other hand, the swap disrupts backbone H-bonding geometry because of the introduction of extra methylene groups into the peptide backbone. In this project, we chose the two-stranded alpha-helical coiled-coil to implement side chain-backbone swap. A pair of 36-residue peptides was designed. The two peptides have identical sequence with four residues in each heptad repeat occupied by glutamyl residues. Each glutamic acid was incorporated either as alpha-glutamyl residue (the peptide is denoted as alpha-Glu-36) or as gamma-glutamyl residue (the peptide is denoted as gamma-Glu-36). The inter-conversion between the two peptides constitutes a side chain-backbone swap. Residues constituting the hydrophobic core of the coiled-coil, however, are left unchanged. The peptide pair was characterized by circular dichroism spectroscopy, reversed-phase liquid chromatography (RPLC), and two-dimensional nuclear magnetic resonance (NMR). The results indicate that alpha-Glu-36 is a two-stranded alpha-helical coiled-coil while gamma-Glu-36 lacks stable structural elements. It is concluded that, at least for coiled-coils where hydrophobic interactions are predominantly long-range, local backbone H-bonding is a required for structural formation, consistent with a hierarchic folding mechanism. The methodological implication of side chain-backbone swap is also discussed.
为了评估主链氢键作用与侧链疏水性在蛋白质结构形成中的相对重要性，提出了一种称为侧链 - 主链交换的方法。这种方法交换某些氨基酸残基（如天冬氨酸、谷氨酸、天冬酰胺、谷氨酰胺、赖氨酸和精氨酸）的侧链和主链部分。这种交换保留了多肽的序列并保持了主链连接的一致性。另一方面，由于在肽主链中引入了额外的亚甲基，这种交换破坏了主链氢键几何结构。在本项目中，我们选择双链α - 螺旋卷曲螺旋来实施侧链 - 主链交换。设计了一对36个残基的肽。这两个肽具有相同的序列，每个七肽重复中有四个残基被谷氨酰残基占据。每个谷氨酸要么作为α - 谷氨酰残基掺入（该肽表示为α - Glu - 36），要么作为γ - 谷氨酰残基掺入（该肽表示为γ - Glu - 36）。这两个肽之间的相互转化构成了侧链 - 主链交换。然而，构成卷曲螺旋疏水核心的残基保持不变。通过圆二色光谱、反相液相色谱（RPLC）和二维核磁共振（NMR）对该肽对进行了表征。结果表明，α - Glu - 36是双链α - 螺旋卷曲螺旋，而γ - Glu - 36缺乏稳定的结构元件。得出的结论是，至少对于疏水相互作用主要是长程的卷曲螺旋来说，局部主链氢键作用是结构形成所必需的，这与分级折叠机制一致。还讨论了侧链 - 主链交换的方法学意义。
