Choe J Y, Fromm H J, Honzatko R B
Department of Biochemistry, Iowa State University, Ames 50011, USA.
Biochemistry. 2000 Jul 25;39(29):8565-74. doi: 10.1021/bi000574g.
Crystal structures of metal-product complexes of fructose 1, 6-bisphosphatase (FBPase) reveal competition between AMP and divalent cations. In the presence of AMP, the Zn(2+)-product and Mg(2+)-product complexes have a divalent cation present only at one of three metal binding sites (site 1). The enzyme is in the T-state conformation with a disordered loop of residues 52-72 (loop 52-72). In the absence of AMP, the enzyme crystallizes in the R-state conformation, with loop 52-72 associated with the active site. In structures without AMP, three metal-binding sites are occupied by Zn(2+) and two of three metal sites (sites 1 and 2) by Mg(2+). Evidently, the association of AMP with FBPase disorders loop 52-72, the consequence of which is the release of cations from two of three metal binding sites. In the Mg(2+) complexes (but not the Zn(2+) complexes), the 1-OH group of fructose 6-phosphate (F6P) coordinates to the metal at site 1 and is oriented for a nucleophilic attack on the bound phosphate molecule. A mechanism is presented for the forward reaction, in which Asp74 and Glu98 together generate a hydroxide anion coordinated to the Mg(2+) at site 2, which then displaces F6P. Development of negative charge on the 1-oxygen of F6P is stabilized by its coordination to the Mg(2+) at site 1.
果糖1,6 - 二磷酸酶（FBPase）的金属产物复合物的晶体结构揭示了AMP与二价阳离子之间的竞争关系。在AMP存在的情况下，Zn(2+) - 产物和Mg(2+) - 产物复合物中，只有三个金属结合位点之一（位点1）存在二价阳离子。该酶处于T态构象，其52 - 72位残基的环（环52 - 72）无序。在没有AMP的情况下，该酶以R态构象结晶，环52 - 72与活性位点相关联。在没有AMP的结构中，三个金属结合位点被Zn(2+)占据，三个金属位点中的两个（位点1和2）被Mg(2+)占据。显然，AMP与FBPase的结合使环52 - 72无序，其结果是三个金属结合位点中的两个释放阳离子。在Mg(2+)复合物（但不是Zn(2+)复合物）中，果糖6 - 磷酸（F6P）的1 - OH基团与位点1的金属配位，并定向对结合的磷酸分子进行亲核攻击。本文提出了正向反应的机制，其中Asp74和Glu98共同产生一个与位点2的Mg(2+)配位的氢氧根阴离子，然后该阴离子取代F6P。F6P的1 - 氧上负电荷的发展通过其与位点1的Mg(2+)配位而得以稳定。
