The Ca2+ ion and membrane binding structure of the Gla domain of Ca-prothrombin fragment 1.
The Ca2+ ion and membrane binding structure of the Gla domain of Ca-prothrombin fragment 1.
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DOI：
10.2210/pdb2pf2/pdb
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发表时间：
1994-01
期刊：
影响因子：
2.9
通讯作者：
M. Soriano-garcia;K. Padmanabhan;A. D. de Vos;A. Tulinsky
中科院分区：
文献类型：
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作者：
M. Soriano-garcia;K. Padmanabhan;A. D. de Vos;A. Tulinsky
Ca-凝血酶原片段 1（凝血酶原残基 1-156）的结构已通过 X 射线晶体学方法以 2.2-A 分辨率解析和细化。在 apo 片段 1 的晶体中，前三分之二的 Gla 结构域（残基 1-48）和两条碳水化合物链（大约 5 kDa）是无序的。当晶体在 Ca2+ 离子存在下生长时，Gla 结构域表现出结合七个 Ca2+ 离子的明确结构，但碳水化合物仍然是无序的。即便如此，晶体 R 因子降低至 0.171。 Gla 结构域的折叠主要由三个不同 α 螺旋的 9-10 转决定。这些匝产生两个由 Gla 侧链组成的内部羧酸盐表面。间隔约 4.0 A 的五个 Ca2+ 离子的聚合物阵列插入到羧酸盐表面之间。 Ca2+离子与Gla羧酸根氧原子和水分子的配位导致在高度复杂的阵列中具有mu-oxo桥的扭曲多面体排列，这很可能协调域的折叠。 Ca2+ 离子相互作用的总体模式是新的，并且不同于迄今为止观察或描述的任何 Ca2+ 离子-蛋白质相互作用。 Ca2+ 离子结合时观察到的荧光猝灭事件是由于二硫键 -π 电子相互作用导致 Gla 结构域的 Trp42 发生 100 度重新定向。 Ca2+ 离子相互作用还提供 N 末端免受乙酰化的保护，因为后者埋在折叠结构中并与 Gla17、Gla21 和 Gla27 形成氢键盐桥。 Gla 结构域及其尾随二硫键单元紧密结合，共同形成结构域样结构。静电势计算表明 Gla 域具有很强的负电性。由于 Gla 残基的大部分羧酸氧原子参与 Ca2+ 离子结合，仅留下少数用于桥接 Ca2+ 离子-磷脂相互作用，因此桥接 Ca2+ 离子的作用通常可能是非特异性的，Ca2+ 离子简单地插入负 Gla 结构域和膜表面的负头基之间。 apo-和Ca-片段1中的kringle结构的折叠本质上是相同的。然而，Gla 结构域的 Ser36-Ala47 螺旋围绕 Cys48 旋转，移动约 30 度，并且螺旋侵入 kringle，产生一些伴随的变化。这些可能与保护携带 Asn101 的碳水化合物免于 Ca 片段 1 结构中的乙酰化有关。
The structure of Ca-prothrombin fragment 1 (residues 1-156 prothrombin) has been solved and refined at 2.2-A resolution by X-ray crystallographic methods. The first two-thirds of the Gla domain (residues 1-48) and two carbohydrate chains (approximately 5 kDa) are disordered in crystals of apo-fragment 1. When crystals are grown in the presence of Ca2+ ions, the Gla domain exhibits a well-defined structure binding seven Ca2+ ions, but the carbohydrate is still disordered. Even so, the crystallographic R factor reduced to 0.171. The folding of the Gla domain is dominated by 9-10 turns of three different alpha-helices. These turns produce two internal carboxylate surfaces composed of Gla side chains. A polymeric array of five Ca2+ ions separated by about 4.0 A intercalates between the carboxylate surfaces. The coordination of the Ca2+ ions with Gla carboxylate oxygen atoms and water molecules leads to distorted polyhedral arrangements with mu-oxo bridges in a highly complex array that most likely orchestrates the folding of the domain. The overall mode of interaction of the Ca2+ ions is new and different from any Ca2+ ion-protein interactions heretofore observed or described. The fluorescence quenching event observed upon Ca2+ ion binding is due to a disulfide-pi-electron interaction that causes a 100 degrees reorientation of Trp42 of the Gla domain. The Ca2+ ion interaction also affords the N-terminus protection from acetylation because the latter is buried in the folded structure and makes hydrogen-bonding salt bridges with Gla17, Gla21, and Gla27. The Gla domain and its trailing disulfide unit associate intimately and together give rise to a domain-like structure. Electrostatic potential calculations indicate that the Gla domain is very electronegative. Since most of the carboxylate oxygen atoms of Gla residues are involved in Ca2+ ion binding, leaving only a few for bridging Ca2+ ion-phospholipid interactions, the role of bridging Ca2+ ions might be generally unspecific, with Ca2+ ions simply intervening between the negative Gla domain and negative head groups of the membrane surface. The folding of the kringle structure in apo- and Ca-fragment 1 is essentially the same. However, the Ser36-Ala47 helix of the Gla domain pivots around Cys48, shifting by approximately 30 degrees, and the helix encroaches on the kringle producing some concomitant changes. These might be related to the protection of carbohydrate carrying Asn101 from acetylation in the Ca-fragment 1 structure.