Development of a high-throughput in vitro assay to identify selective inhibitors for human ALDH1A1.
Development of a high-throughput in vitro assay to identify selective inhibitors for human ALDH1A1.
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DOI：
10.1016/j.cbi.2014.10.028
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发表时间：
2015-06-05
影响因子：
5.1
通讯作者：
Hurley, Thomas D.
中科院分区：
文献类型：
--
作者：
Morgan, Cynthia A.;Hurley, Thomas D.
关键词：
人醛脱氢酶(ALDH)超家族由至少19种代谢内源和外源醛的酶组成。目前，还没有针对ALDH1A1的商业上可用的抑制剂，但对结构和功能相似的ALDH2几乎没有影响。在这里，我们提出了第一个人类ALDH1A1结构，作为脱辅酶，并与其辅因子NADH形成复合体，分辨率分别为1.75？和2.1？ALDH1A1与其他密切相关的ALDH酶的辅因子结合位点的结构比较表明，具有高度的相似性。为了最大限度地减少通过干扰这两种同工酶保守的辅因子结合位点而抑制它们的化合物的发现，本研究报告了使用体外、NAD+独立的、基于酯酶的64,000种化合物的高通量筛选(HTS)来发现新型的、选择性的ALDH1A1抑制剂。我们描述了256个改变ALDH1A1酯酶活性的点击。进一步分析了67个化合物对乙醛氧化的影响，其中30个化合物比ALDH2和ALDH3A1选择性地抑制ALDH1A1。一种化合物抑制ALDH1A1和ALDH2，而另一种化合物抑制ALDH1A1、ALDH2和更远亲的ALDH3A1。结果表明，这种体外酶活性筛选方法成功地鉴定了具有高度同工酶选择性的ALDH1A1抑制剂。通过这一筛选鉴定的化合物加上筛选方法本身代表了开发高效和选择性的ALDH1A1抑制剂的起点，可用于更好地了解这种酶在正常和疾病状态下的作用。
The human aldehyde dehydrogenase (ALDH) superfamily consists of at least 19 enzymes that metabolize endogenous and exogenous aldehydes. Currently, there are no commercially available inhibitors that target ALDH1A1 but have little to no effect on the structurally and functionally similar ALDH2. Here we present the first human ALDH1A1 structure, as the apoenzyme and in complex with its cofactor NADH to a resolution of 1.75 Å and 2.1 Å, respectfully. Structural comparisons of the cofactor binding sites in ALDH1A1 with other closely related ALDH enzymes illustrate a high degree of similarity. In order to minimize discovery of compounds that inhibit both isoenzymes by interfering with their conserved cofactor binding sites, this study reports the use of an in vitro, NAD+-independent, esterase-based high-throughput screen (HTS) of 64,000 compounds to discover novel, selective inhibitors of ALDH1A1. We describe 256 hits that alter the esterase activity of ALDH1A1. The effects on aldehyde oxidation of 67 compounds were further analyzed, with 30 selectively inhibiting ALDH1A1 compared to ALDH2 and ALDH3A1. One compound inhibited ALDH1A1 and ALDH2, while another inhibited ALDH1A1, ALDH2, and the more distantly related ALDH3A1. The results presented here indicate that this in vitro enzyme activity screening protocol successfully identified ALDH1A1 inhibitors with a high degree of isoenzyme selectivity. The compounds identified via this screen plus the screening methodology itself represent a starting point for the development of highly potent and selective inhibitors of ALDH1A1 that may be utilized to better understand the role of this enzyme in both normal and disease states.
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影响因子：
5.1
作者：
Moreb, Jan S.;Ucar, Deniz;Han, Shuhong;Amory, John K.;Goldstein, Alex S.;Ostmark, Blanca;Chang, Lung-Ji
通讯作者：
Chang, Lung-Ji
影响因子：
3.2
作者：
Parajuli, Bibek;Georgiadis, Taxiarchis M.;Fishel, Melissa L.;Hurley, Thomas D.
通讯作者：
Hurley, Thomas D.
DOI：
10.1107/s0907444904019158
发表时间：
2004-12-01
影响因子：
2.2
作者：
Emsley, P;Cowtan, K
通讯作者：
Cowtan, K
DOI：
10.1038/nsb0497-317
发表时间：
1997-04-01
期刊：
NATURE STRUCTURAL BIOLOGY
影响因子：
--
作者：
Liu, ZJ;Sun, YJ;Wang, BC
通讯作者：
Wang, BC
影响因子：
4.8
作者：
Khanna, May;Chen, Che-Hong;Hurley, Thomas D.
通讯作者：
Hurley, Thomas D.