THE ROLE OF INTERHELICAL IONIC INTERACTIONS IN CONTROLLING PROTEIN-FOLDING AND STABILITY - DE-NOVO DESIGNED SYNTHETIC 2-STRANDED ALPHA-HELICAL COILED-COILS

被引:204
作者
ZHOU, NE
KAY, CM
HODGES, RS
机构
[1] UNIV ALBERTA, DEPT BIOCHEM, EDMONTON T6G 2S2, AB, CANADA
[2] UNIV ALBERTA, PROT ENGN NETWORK CTR EXCELLENCE, EDMONTON T6G 2S2, AB, CANADA
关键词
PROTEIN DE-NOVO DESIGN; COILED-COILS; LEUCINE ZIPPER; ALPHA-HELICES; INTERHELICAL IONIC INTERACTIONS;
D O I
10.1006/jmbi.1994.1250
中图分类号
Q5 [生物化学]; Q7 [分子生物学];
学科分类号
071010 ; 081704 ;
摘要
The role of interchain ionic interactions in controlling protein folding and stability has been studied by using de novo designed synthetic two-stranded α-helical coiled-coils. The model coiled-coil (denoted as EK) consists of two identical 35-residue polypeptide chains with a heptad repeat KgLaGbAcLdEeKf and a Cys residue at position 2 and an Ala residue at position 16 in each chain. The Lys residues at positions "g" in one chain and Glu residues at positions "c" in the other chain are expected to form interchain ion-pairs in the coiled-coil structure. This peptide forms a stable coiled-coil structure in benign medium (50 mM KCl, 25 mM PO4, pH7) with a [urea]1/2 value of 3.5 M. In contrast, two peptide analogs EE (EgLaGbAcLdEeKf) and KK (KgLaGbAcLdKeEf), which differ from EK in that EE contains only negatively charged Glu residues and KK contains only positively charged Lys residues at both positions e and g, each show a random coil structure in benign buffer. However, peptide EE or KK can form a stable coiled-coil structure if the interchain ionic repulsions are effectively suppressed either by changing pH or by using high salt concentrations. An equimolar mixture of these two peptides displays 100% α-helical content under the same conditions. These results demonstrate that although the interhelical ionic attractions are not essential for coiled-coil formation, a large number of these weak interactions can play an important role in the assembly of helices. Though interhelical ionic repulsions destabilize the homo-stranded coiled-coil, electrostatic attractions may stabilize the hetero-stranded coiled-coil. In addition, this study also suggests that the folding process for these synthetic model coiled-coils does not involve a single-stranded α-helix as a significantly populated folding intermediate.
引用
收藏
页码:500 / 512
页数:13
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