Characterization of AKR1B16, a novel mouse aldo-keto reductase
Characterization of AKR1B16, a novel mouse aldo-keto reductase
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DOI：
10.1016/j.cbi.2017.03.007
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发表时间：
2017-10-01
影响因子：
5.1
通讯作者：
Farres, Jaume
中科院分区：
文献类型：
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作者：
Gimenez-Dejoz, Joan;Weber, Susanne;Farres, Jaume
醛酮还原酶（Aldo-keto reductases，AKR）在哺乳动物中分布于三个家族和多个亚家族。小鼠Akr 1b 3基因明显与人AKR 1B 1基因同源，两者都编码醛糖还原酶，它们的基因产物显示出相似的组织分布、渗透压调节和动力学特性。相比之下，在啮齿类动物中尚未鉴定出人AKR 1B 10和AKR1B15.1的明确直系同源物。虽然在小鼠中已经鉴定和表征了另外两种AKR，AKR 1B 7和AKR 1B 8，但是它们似乎都没有表现出与人AKR相似的特性。最近，一种新的小鼠AKR基因Akr 1b 16被注释，并且相应的基因产物AKR 1B 16（分别与AKR 1B 10和AKR1B15.1共享83%和80%的氨基酸序列同一性）在细菌表达系统中表达为不溶性和无活性的蛋白。在这里，我们描述了表达和纯化的可溶性和酶活性的AKR 1B 16从E。大肠杆菌中使用三个分子伴侣系统。AKR 1B 16的结构模型允许估计其活性位点口袋体积，其比AKR 1B 10（279埃（3））和AKR1B15.1（60埃（3））宽得多（402埃（3））。AKR 1B 16使用NADPH作为辅因子还原脂肪族和芳香族羰基化合物，具有中等或低活性（最高kcat值约为5 min（-1））。该酶的最佳底物为吡啶-3-醛。AKR 1B 16与经典的AKR抑制剂一起显示出较差的抑制，托司他是最有效的。动力学和抑制特性与大鼠AKR 1B 17相似，但与人类酶不同。此外，AKR 1B 16以NADP（+）依赖性方式催化17 b-羟基类固醇的氧化。这些结果与系统发育分析一起表明，小鼠AKR 1B 16是大鼠AKR 1B 17的直系同源物，但不是人AKR 1B 10或AKR1B15.1的直系同源物。这些人酶在鼠物种中没有对应物，这通过在系统发育树中形成单独的簇以及它们与视黄醇的独特活性来证明。(C)2017爱思唯尔B. V.保留所有权利。
Aldo-keto reductases (AKRs) are distributed in three families and multiple subfamilies in mammals. The mouse Akr1b3 gene is clearly orthologous to human AKR1B1, both coding for aldose reductase, and their gene products show similar tissue distribution, regulation by osmotic stress and kinetic properties. In contrast, no unambiguous orthologs of human AKR1B10 and AKR1B15.1 have been identified in rodents. Although two more AKRs, AKR1B7 and AKR1B8, have been identified and characterized in mouse, none of them seems to exhibit properties similar to the human AKRs. Recently, a novel mouse AKR gene, Akr1b16, was annotated and the respective gene product, AKR1B16 (sharing 83% and 80% amino acid sequence identity with AKR1B10 and AKR1B15.1, respectively), was expressed as insoluble and inactive protein in a bacterial expression system. Here we describe the expression and purification of a soluble and enzymatically active AKR1B16 from E. coli using three chaperone systems. A structural model of AKR1B16 allowed the estimation of its active-site pocket volume, which was much wider (402 angstrom(3)) than those of AKR1B10 (279 angstrom(3)) and AKR1B15.1 (60 angstrom(3)). AKR1B16 reduced aliphatic and aromatic carbonyl compounds, using NADPH as a cofactor, with moderate or low activity (highest kcat values around 5 min(-1)). The best substrate for the enzyme was pyridine-3-aldehyde. AKR1B16 showed poor inhibition with classical AKR inhibitors, tolrestat being the most potent. Kinetics and inhibition properties resemble those of rat AKR1B17 but differ from those of the human enzymes. In addition, AKR1B16 catalyzed the oxidation of 17b-hydroxysteroids in a NADP(+)-dependent manner. These results, together with a phylogenetic analysis, suggest that mouse AKR1B16 is an ortholog of rat AKR1B17, but not of human AKR1B10 or AKR1B15.1. These human enzymes have no counterpart in the murine species, which is evidenced by forming a separate cluster in the phylogenetic tree and by their unique activity with retinaldehyde. (C) 2017 Elsevier B.V. All rights reserved.