Lin Hua-Yang, Liu Jyung-Hurng, Cheng Ka-Lik, Lin Jia-Yun, Liu Ni-Rung, Meng Menghsiao
Graduate Institute of Biotechnology, National Chung Hsing University (NCHU), 250 Kuo-Kuang Road, Taichung, Taiwan 40227.
Graduate Institute of Genomics and Bioinformatics, NCHU, Taichung, Taiwan 40227.
Biochem Biophys Rep. 2015 Apr 30;2:14-22. doi: 10.1016/j.bbrep.2015.04.003. eCollection 2015 Jul.
Phosphoglucose isomerase (PGI) catalyzes the interconversion between glucose 6-phosphate and fructose 6-phosphate in the glycolysis pathway. In mammals, the enzyme is also identical to the extracellular proteins neuroleukin, tumor-secreted autocrine motility factor (AMF) and differentiation and maturation mediator for myeloid leukemia. Hereditary deficiency of the enzyme causes non-spherocytic hemolytic anemia in human. In the present study, a novel interaction between GTP and human PGI was corroborated by UV-induced crosslinking, affinity purification and kinetic study. GTP not only inhibits the isomerization activity but also compromises the AMF function of the enzyme. Kinetic studies, including the Yonetani-Theorell method, suggest that GTP is a competitive inhibitor with a value of 63 μM and the GTP-binding site partially overlaps with the catalytic site. In addition, GTP stabilizes the structure of human PGI against heat- and detergent-induced denaturation. Molecular modelling and dynamic simulation suggest that GTP is bound in a -conformation with the γ-phosphate group located near the phosphate-binding loop and the ribose moiety positioned away from the active-site residues.
磷酸葡萄糖异构酶（PGI）在糖酵解途径中催化6-磷酸葡萄糖和6-磷酸果糖之间的相互转化。在哺乳动物中，该酶还等同于细胞外蛋白神经白细胞素、肿瘤分泌的自分泌运动因子（AMF）以及髓系白血病的分化和成熟介质。该酶的遗传性缺乏会导致人类非球形细胞溶血性贫血。在本研究中，通过紫外线诱导交联、亲和纯化和动力学研究证实了GTP与人类PGI之间存在一种新的相互作用。GTP不仅抑制异构化活性，还损害该酶的AMF功能。包括米氏-泰罗尔方法在内的动力学研究表明，GTP是一种竞争性抑制剂，其 值为63 μM，且GTP结合位点与催化位点部分重叠。此外，GTP可稳定人类PGI的结构，使其抵抗热和去污剂诱导的变性。分子建模和动态模拟表明，GTP以 -构象结合，γ-磷酸基团位于磷酸结合环附近，核糖部分远离活性位点残基。
