Development of A Continuous Fluorescence-Based Assay for N-Terminal Acetyltransferase D.
Development of A Continuous Fluorescence-Based Assay for N-Terminal Acetyltransferase D.
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DOI：
10.3390/ijms22020594
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发表时间：
2021-01-08
影响因子：
5.6
通讯作者：
Huang R
中科院分区：
文献类型：
--
作者：
Ho YH;Chen L;Huang R
N-末端乙酰基转移酶(NAT)催化的N-末端乙酰化在蛋白质调节中具有多种生物学功能。N-末端乙酰转移酶D(NatD)是一种最具特异性的NAT，仅以组蛋白H4和H_2A蛋白为底物。NATD的调节失调与结直肠癌和肺癌的进展有关，这意味着它在癌症中的治疗潜力。然而，目前还没有关于NATD的抑制剂的报道。为了促进小分子NATD抑制剂的发现，我们报道了一种基于荧光的乙酰基转移酶检测方法，该方法以384孔高通量筛选(HTS)格式为基础，通过监测辅酶A的形成来产生荧光信号。该方法的Z‘因子为0.77，变异系数为6%，是一种可靠的HTS检测方法。对1280种药理活性化合物的初步筛选和随后的验证确定了两项命中，证实了这种荧光分析在HTS中的应用。
N-terminal acetylation catalyzed by N-terminal acetyltransferases (NATs) has various biological functions in protein regulation. N-terminal acetyltransferase D (NatD) is one of the most specific NAT with only histone H4 and H2A proteins as the known substrates. Dysregulation of NatD has been implicated in colorectal and lung cancer progression, implying its therapeutic potential in cancers. However, there is no reported inhibitor for NatD yet. To facilitate the discovery of small-molecule NatD inhibitors, we report the development of a fluorescence-based acetyltransferase assay in 384-well high-throughput screening (HTS) format through monitoring the formation of coenzyme A. The fluorescent signal is generated from the adduct in the reaction between coenzyme A and fluorescent probe ThioGlo4. The assay exhibited a Z′-factor of 0.77 and a coefficient of variation of 6%, indicating it is a robust assay for HTS. A pilot screen of 1280 pharmacologically active compounds and subsequent validation identified two hits, confirming the application of this fluorescence assay in HTS.
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DOI：
10.1007/978-1-4939-6850-3_2
发表时间：
2017-01-01
期刊：
PROTEIN TERMINAL PROFILING: METHODS AND PROTOCOLS
影响因子：
--
作者：
Foyn, Havard;Thompson, Paul R.;Arnesen, Thomas
通讯作者：
Arnesen, Thomas
NatD 通过阻止组蛋白 H4 丝氨酸磷酸化激活 Slug 表达来促进肺癌进展
影响因子：
16.6
作者：
Ju J;Chen A;Deng Y;Liu M;Wang Y;Wang Y;Nie M;Wang C;Ding H;Yao B;Gui T;Li X;Xu Z;Ma C;Song Y;Kvansakul M;Zen K;Zhang CY;Luo C;Fang M;Huang DCS;Allis CD;Tan R;Zeng CK;Wei J;Zhao Q
通讯作者：
Zhao Q
影响因子：
9.8
作者：
Arnesen T
通讯作者：
Arnesen T
影响因子：
4.8
作者：
Richardson, Stacie L.;Mao, Yunfei;Huang, Rong
通讯作者：
Huang, Rong
DOI：
10.1007/978-1-4939-6850-3_1
发表时间：
2017-01-01
期刊：
PROTEIN TERMINAL PROFILING: METHODS AND PROTOCOLS
影响因子：
--
作者：
Drazic, Adrian;Arnesen, Thomas
通讯作者：
Arnesen, Thomas