Lee Ji Hyun, Jeffery Constance J
Laboratory for Molecular Biology, Department of Biological Sciences, MC567, 900 Ashland Ave., University of Illinois at Chicago, Chicago, IL 60607, USA.
Protein Sci. 2005 Mar;14(3):727-34. doi: 10.1110/ps.041070205. Epub 2005 Feb 2.
Phosphoglucose isomerase (PGI) catalyzes the isomerization of D-glucose-6-phosphate (G6P) and D-fructose-6-phosphate (F6P) in glycolysis and gluconeogenesis. Analysis of previously reported X-ray crystal structures of PGI without ligand, with the cyclic form of F6P, or with inhibitors that mimic the cis-enediol intermediate led to proposed mechanisms for the ring opening and isomerization steps in the multistep catalytic mechanism. To help complete our model of the overall mechanism, information is needed about the state of PGI between the ring opening and isomerization steps, in other words, a structure of the enzyme complexed with the open form of a substrate or an analog. Here, we report the crystal structure of rabbit PGI complexed with D-sorbitol-6-phosphate (S6P), an analog of the open chain form of G6P, at 2.0 A resolution. As was seen in the PGI/F6P structure, a helix containing amino acid residues 512-520 is found in the "out" position, which provides sufficient space in the active site for a substrate in its cyclic form and which is probably the location of that helix just after ring opening (or just before ring closure). However, the S6P ligand is in an extended conformation, as was seen previously with ligands that mimic the cis-enediol intermediate. The extended conformation enables the ligand to interact with Glu357, which transfers a proton during the isomerization step. The PGI/S6P structure represents the conformation of the enzyme and substrate between the ring opening (or ring closing) step and the isomerization step and helps to complete the model for PGI's catalytic mechanism.
磷酸葡萄糖异构酶(PGI)在糖酵解和糖异生过程中催化6-磷酸-D-葡萄糖(G6P)和6-磷酸-D-果糖(F6P)的异构化反应。对先前报道的无配体、与F6P环状形式结合或与模拟顺式烯二醇中间体的抑制剂结合的PGI的X射线晶体结构进行分析,得出了多步催化机制中环开环和异构化步骤的推测机制。为了完善我们的整体机制模型,需要有关PGI在开环和异构化步骤之间状态的信息,换句话说,需要一种与底物或类似物的开放形式复合的酶的结构。在此,我们报道了兔PGI与6-磷酸-D-山梨醇(S6P,G6P开链形式的类似物)复合的晶体结构,分辨率为2.0埃。正如在PGI/F6P结构中所见,包含氨基酸残基512-520的螺旋处于“外部”位置,这为环状形式的底物在活性位点提供了足够的空间,并且这可能是开环后(或闭环前)该螺旋的位置。然而,S6P配体处于伸展构象,正如先前与模拟顺式烯二醇中间体的配体所见。伸展构象使配体能够与Glu357相互作用,Glu357在异构化步骤中转移一个质子。PGI/S6P结构代表了开环(或闭环)步骤和异构化步骤之间酶和底物的构象,并有助于完善PGI催化机制的模型。