Bouckaert J, Poortmans F, Wyns L, Loris R
Laboratorium voor Ultrastructuur, Vlaams Interuniversitair Instituut voor Biotechnologie, Vrije Universiteit Brussel and Vlaamse Instelling voor Technologisch Onderzoek, Boeretang 200, B-2400 Mol, Belgium.
J Biol Chem. 1996 Jul 5;271(27):16144-50. doi: 10.1074/jbc.271.27.16144.
The lectin concanavalin A (ConA) sequentially binds a transition metal ion in the metal-binding site S1 and a calcium ion in the metal-binding site S2 to form its saccharide-binding site. Metal-free ConA crystals soaked with either Zn2+ (apoZn-ConA) or Co2+ (apoCo-ConA) display partial binding of these ions in the proto-transition metal-binding site, but no further conformational changes are observed. These structures can represent the very first step in going from metal-free ConA toward the holoprotein. In the co-crystals of metal-free ConA with Zn2+ (Zn-ConA), the zinc ion can fully occupy the S1 site. The positions of the carboxylate ligands Asp10 and Asp19 that bridge the S1 and S2 sites are affected. The ligation to Zn2+ orients Asp10 optimally for calcium ligation and stabilizes Asp19 by a hydrogen bond to one of its water ligands. The neutralizing and stabilizing effect of the binding of Zn2+ in S1 is necessary to allow for subsequent Ca2+ binding in the S2 site. However, the S2 site of monometallized ConA is still disrupted. The co-crystals of metal-free ConA with both Zn2+ and Ca2+ contain the active holoprotein (ConA ZnCa). Ca2+ has induced large conformational changes to stabilize its hepta-coordination in the S2 site, which comprise the trans to cis isomerization of the Ala207-Asp208 peptide bond accompanied by the formation of the saccharide-binding site. The Zn2+ ligation in ConA ZnCa is similar to Mn2+, Cd2+, Co2+, or Ni2+ ligation in the S1 site, in disagreement with earlier extended x-ray absorption fine structure results that suggested a lower coordination number for Zn2+.
凝集素伴刀豆球蛋白A（ConA）依次在金属结合位点S1结合一个过渡金属离子，并在金属结合位点S2结合一个钙离子，以形成其糖结合位点。用Zn2+（无金属锌-ConA）或Co2+（无金属钴-ConA）浸泡的无金属ConA晶体在原过渡金属结合位点显示这些离子的部分结合，但未观察到进一步的构象变化。这些结构可以代表从无金属ConA向全蛋白转变的第一步。在无金属ConA与Zn2+的共晶体（锌-ConA）中，锌离子可以完全占据S1位点。连接S1和S2位点的羧酸盐配体天冬氨酸10（Asp10）和天冬氨酸19（Asp19）的位置受到影响。与Zn2+的连接使Asp10以最佳方向进行钙连接，并通过与其一水配体形成氢键来稳定Asp19。S1中Zn2+结合的中和和稳定作用对于随后S2位点中Ca2+的结合是必要的。然而，单金属化ConA的S2位点仍然受到破坏。无金属ConA与Zn2+和Ca2+的共晶体包含活性全蛋白（ConA ZnCa）。Ca2+诱导了大的构象变化，以稳定其在S2位点的七配位，这包括Ala207-Asp208肽键从反式到顺式的异构化，并伴随着糖结合位点的形成。ConA ZnCa中Zn2+的配位与S1位点中Mn2+、Cd2+、Co2+或Ni2+的配位相似，这与早期扩展X射线吸收精细结构结果不一致，早期结果表明Zn2+的配位数较低。
