Dieckmann G R, McRorie D K, Lear J D, Sharp K A, DeGrado W F, Pecoraro V L
Department of Chemistry, University of Michigan, 930 North University, Ann Arbor, MI 48109-1055, USA.
J Mol Biol. 1998 Jul 31;280(5):897-912. doi: 10.1006/jmbi.1998.1891.
To define the delicate interplay between metal chelation, protein folding and function in metalloproteins, a family of de novo-designed peptides was synthesized that self-assemble in aqueous solution to form two and three-stranded alpha-helical coiled coils. Each peptide contains a single Cys residue at an a or d position of the heptad repeat. Peptide association thus produces a Cys-rich coordination environment that has been used to bind Hg(II) ions. These peptides display a pH-dependent association, with trimers observed above the pKa of Glu side-chains and dimers below this value. Finite-difference Poisson-Boltzmann calculations suggest that the dimeric state decreases the unfavorable electrostatic interactions between positively charged Lys side-chains (relative to the trimer). The Cys-containing peptides bind Hg(II) in a position-dependent fashion. Cys at a positions form three-coordinate Hg complexes at high pH where the trimeric aggregation state predominates, and two-coordinate complexes at lower pH. A d position Cys, however, is only able to generate the two-coordinate complex, illustrating the difference in coordination geometry between the two positions in the coiled coil. The binding of Hg(II) was also shown to substantially increase the stability of the helical aggregates.
为了确定金属螯合、蛋白质折叠与金属蛋白功能之间微妙的相互作用，合成了一族从头设计的肽，它们在水溶液中自组装形成双股和三股α-螺旋卷曲螺旋。每个肽在七肽重复序列的a或d位置含有一个半胱氨酸（Cys）残基。肽的缔合因此产生了一个富含半胱氨酸的配位环境，该环境已被用于结合Hg(II)离子。这些肽表现出pH依赖性缔合，在谷氨酸（Glu）侧链的pKa以上观察到三聚体，在此值以下观察到二聚体。有限差分泊松-玻尔兹曼计算表明，二聚体状态减少了带正电荷的赖氨酸（Lys）侧链之间不利的静电相互作用（相对于三聚体）。含半胱氨酸的肽以位置依赖的方式结合Hg(II)。a位置的半胱氨酸在高pH下形成三配位Hg配合物，此时三聚体聚集状态占主导，在较低pH下形成二配位配合物。然而，d位置的半胱氨酸只能生成二配位配合物，这说明了卷曲螺旋中两个位置之间配位几何结构的差异。还表明Hg(II)的结合显著增加了螺旋聚集体的稳定性。
