Hill C P, Dauter Z, Dodson E J, Dodson G G, Dunn M F
Department of Chemistry, University of York, Heslington, U.K.
Biochemistry. 1991 Jan 29;30(4):917-24. doi: 10.1021/bi00218a006.
Metal ion binding to the insulin hexamer has been investigated by crystallographic analysis. Cadmium, lead, and metal-free hexamers have been refined to R values of 0.181, 0.172, and 0.172, against data of 1.9-, 2.5-, and 2.5-A resolution, respectively. These structures have been compared with each other and with the isomorphous two-zinc insulin. The structure of the metal-free hexamer shows that the His(B10) imidazole rings are arranged in a preformed site that binds a water molecule and is poised for Zn2+ coordination. The structure of the cadmium derivative shows that the binding of Cd2+ at the center of the hexamer is unusual. There are three symmetry-related sites located within 2.7 A of each other, and this position is evidently one-third occupied. It is also shown that the coordinating B13 glutamate side chains of this derivative have two partially occupied conformations. One of these conformations is two-thirds occupied and is very similar to that seen in two-zinc insulin. The other, one-third-occupied conformation, is seen to coordinate the one-third-occupied metal ion. The binding of Ca2+ to insulin is assumed to be essentially identical with that of Cd2+. Thus, we conclude that the Ca2+ binding site in the insulin hexamer is unlike that of any other known calcium binding protein. The crystal structures reported herein explain how binding of metal ions stabilizes the insulin hexamer. The role of metal ions in hexamer assembly and dissociation is discussed.
通过晶体学分析研究了金属离子与胰岛素六聚体的结合。镉、铅和无金属六聚体分别针对分辨率为1.9 Å、2.5 Å和2.5 Å的数据精修至R值为0.181、0.172和0.172。已将这些结构相互比较，并与同晶型的双锌胰岛素进行了比较。无金属六聚体的结构表明，His(B10)咪唑环排列在一个预先形成的位点，该位点结合一个水分子并为Zn2+配位做好准备。镉衍生物的结构表明，Cd2+在六聚体中心的结合方式不同寻常。有三个对称相关的位点，彼此距离在2.7 Å以内，且该位置显然被占据了三分之一。还表明，该衍生物的配位B13谷氨酸侧链有两种部分占据的构象。其中一种构象被占据了三分之二，与双锌胰岛素中的构象非常相似。另一种构象被占据了三分之一，可配位三分之一被占据的金属离子。假定Ca2+与胰岛素的结合与Cd2+基本相同。因此，我们得出结论，胰岛素六聚体中的Ca2+结合位点与任何其他已知的钙结合蛋白的位点不同。本文报道的晶体结构解释了金属离子的结合如何稳定胰岛素六聚体。讨论了金属离子在六聚体组装和解离中的作用。
