Allosteric kinetics of the isoform 1 of human glucosamine-6-phosphate deaminase
Allosteric kinetics of the isoform 1 of human glucosamine-6-phosphate deaminase
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DOI：
10.1016/j.bbapap.2011.07.010
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发表时间：
2011-12-01
影响因子：
3.2
通讯作者：
Calcagno, Mario L.
中科院分区：
文献类型：
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作者：
Alvarez-Anorve, Laura I.;Alonzo, Diego A.;Calcagno, Mario L.
人类基因组包含两个编码葡萄糖胺-6-磷酸脱氨酶（GNPDA，EC 3.5.99.6）两种亚型的基因。同工型 1 已从多种动物来源中纯化，并且以 1.75 埃的分辨率解析了人类重组酶的晶体结构（PDB ID：1NE7）。尽管人类和大肠杆菌 GNPDA 的结构非常相似，但它们的变构动力学表现出显着差异。变构位点配体 N-乙酰氨基葡萄糖 6-磷酸 (GlcNAc6P) 是大肠杆菌 GNPDA K 型激活剂，对 hGNPDA1 具有不寻常的混合变构效应，表现为 V 激活剂和 K 抑制剂（抗过敏或交叉混合 K-V+ 效应）。在不存在 GlcNAc6P 的情况下，酶的表观 k(cat) 非常低，以至于 GlcNAc6P 充当必需的激活剂。此外，还观察到底物抑制取决于 GlcNAc6P 浓度。所有这些动力学特性都可以在带有一些附加假设的莫诺变构模型框架内得到很好的描述。这些不寻常的动力学特性表明，hGNPDA1 对于维持 UDP-GlcNAc6P 池的足够水平可能很重要，UDP-GlcNAc6P 是细胞中许多反应的 N-乙酰氨基葡萄糖供体。在本研究中，我们还通过构建和研究在位置 268 和 275 截短的两个突变体，探索了 hGNPDA1 酶 C 端延伸（亚型 2 中不存在）的可能功能意义。(C) 2011 Elsevier B.V. 保留所有权利。
The human genome contains two genes encoding for two isoforms of the enzyme glucosamine-6-phosphate deaminase (GNPDA, EC 3.5.99.6). Isoform 1 has been purified from several animal sources and the crystallographic structure of the human recombinant enzyme was solved at 1.75 angstrom resolution (PDB ID: 1NE7). In spite of their great structural similarity, human and Escherichia coli GNPDAs show marked differences in their allosteric kinetics. The allosteric site ligand, N-acetylglucosamine 6-phosphate (GlcNAc6P), which is an activator of the K-type of E. coli GNPDA has an unusual mixed allosteric effect on hGNPDA1, behaving as a V activator and a K inhibitor (antiergistic or crossed mixed K-V+ effect). In the absence of GlcNAc6P, the apparent k(cat) of the enzyme is so low, that GlcNAc6P behaves as an essential activator. Additionally, substrate inhibition, dependent on GIcNAc6P concentration, is observed. All these kinetic properties can be well described within the framework of the Monod allosteric model with some additional postulates. These unusual kinetic properties suggest that hGNPDA1 could be important for the maintenance of an adequate level of the pool of the UDP-GlcNAc6P, the N-acetylglucosylaminyl donor for many reactions in the cell. In this research we have also explored the possible functional significance of the C-terminal extension of hGNPDA1 enzyme, which is not present in isoform 2, by constructing and studying two mutants truncated at positions 268 and 275. (C) 2011 Elsevier B.V. All rights reserved.