Name: capturing the interaction kinetics of an ion channel protein with small molecules by the bio-layer interferometry assay
Uploaded: 2018-03-07T13:30:00
Description: Watch this Scientific Journal Video about Capturing the Interaction Kinetics of an Ion Channel Protein with Small Molecules by the Bio-layer Interferometry Assay at JoVE.com

这项工作得到了生物 ID 中心和上海交通大学交叉学科研究基金 (YG2016QN66)、中国国家自然科学基金 (31271217) 和中国国家基础研究项目 (2014CB910304) 的支持。

注: 连续表达标记标记的 hEAG1 通道蛋白的 HEK-293T 细胞系由转染 pCDH 慢病毒载体质粒组成, 其中含有 hEAG1 的 DNA 序列, 在远端 C 终点有一条旗, 随后为 HEK-293T 细胞。嘌呤霉素的选择, 如前面所描述的15。
1. HEK-293T 细胞标记标记 hEAG1 通道蛋白的亲和纯化
<ol><li>解冻细胞稳定地表达 hEAG1 渠道从液氮入温暖的水 (37 °c) 快速地。将细胞 (大约 5 x 106冷冻细胞) ?...

<ol>
	<li>Peetla, C., Stine, A., Labhasetwar, V. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Biophysical+Interactions+with+Model+Lipid+Membranes:+Applications+in+Drug+Discovery+and+Drug+Delivery.">Biophysical Interactions with Model Lipid Membranes: Applications in Drug Discovery and Drug Delivery.</a> Mol. Pharm. 6 (5), 1264-1276 (2009).</li><li>van de Waterbeemd, H., Smith, D. A., Beaumont, K., Walker, D. K. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Property-based+design:+Optimization+of+drug+absorption+and+pharmacokinetics.">Property-based design: Optimization of drug absorption and pharmacokinetics.</a> J. Med. Chem. 44 (9), 1313-1333 (2001).</li><li>Papo, N., Shai, Y. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Exploring+peptide+membrane+interaction+using+surface+plasmon+resonance:+Differentiation+between+pore+formation+versus+membrane+disruption+by+lytic+peptides.">Exploring peptide membrane interaction using surface plasmon resonance: Differentiation between pore formation versus membrane disruption by lytic peptides.</a> Biochemistry. 42 (2), 458-466 (2003).</li><li>Wienken, C. J., Baaske, P., Rothbauer, U., Braun, D., Duhr, S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Protein-binding+assays+in+biological+liquids+using+microscale+thermophoresis.">Protein-binding assays in biological liquids using microscale thermophoresis.</a> Nat. Commun. 1, 100-106 (2010).</li><li>Fechner, P., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Size+does+matter!+Label-free+detection+of+small+molecule-protein+interaction.">Size does matter! Label-free detection of small molecule-protein interaction.</a> Anal. Bioanal. Chem. 406 (17), 4033-4051 (2014).</li><li>Wallner, J., Lhota, G., Jeschek, D., Mader, A., Vorauer-Uhl, K. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Application+of+Bio-Layer+Interferometry+for+the+analysis+of+protein/liposome+interactions.">Application of Bio-Layer Interferometry for the analysis of protein/liposome interactions.</a> J. Pharm. Biomed. Anal. 72, 150-154 (2013).</li><li>Wartchow, C. A., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Biosensor-based+small+molecule+fragment+screening+with+biolayer+interferometry.">Biosensor-based small molecule fragment screening with biolayer interferometry.</a> J. Comput. Aided Mol. Des. 25 (7), 669-676 (2011).</li><li>Ekiert, D. C., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+Highly+Conserved+Neutralizing+Epitope+on+Group+2+Influenza+A+Viruses.">A Highly Conserved Neutralizing Epitope on Group 2 Influenza A Viruses.</a> Science. 333 (6044), 843-850 (2011).</li><li>Shah, N. B., Duncan, T. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Bio-layer+Interferometry+for+Measuring+Kinetics+of+Protein-protein+Interactions+and+Allosteric+Ligand+Effects.">Bio-layer Interferometry for Measuring Kinetics of Protein-protein Interactions and Allosteric Ligand Effects.</a> J. Vis. Exp. (84), e51383 (2014).</li><li>Occhiodoro, T., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Cloning+of+a+human+ether-a-go-go+potassium+channel+expressed+in+myoblasts+at+the+onset+of+fusion.">Cloning of a human ether-a-go-go potassium channel expressed in myoblasts at the onset of fusion.</a> Febs Lett. 434 (1-2), 177-182 (1988).</li><li>Pardo, L. A., Stuhmer, W. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Eag1:+An+emerging+oncological+target.">Eag1: An emerging oncological target.</a> Cancer. Res. 68 (6), 1611-1613 (2008).</li><li>Simons, C., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Mutations+in+the+voltage-gated+potassium+channel+gene+KCNH1+cause+Temple-Baraitser+syndrome+and+epilepsy.">Mutations in the voltage-gated potassium channel gene KCNH1 cause Temple-Baraitser syndrome and epilepsy.</a> Nat. Genet. 47 (1), 73-77 (2015).</li><li>Kortum, F., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Mutations+in+KCNH1+and+ATP6V1B2+cause+Zimmermann-Laband+syndrome.">Mutations in KCNH1 and ATP6V1B2 cause Zimmermann-Laband syndrome.</a> Nat. Genet. 47 (6), 661-667 (2015).</li><li>Han, B., Tokay, T., Zhang, G. M., Sun, P., Hou, S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Eag1+K++Channel:+Endogenous+Regulation+and+Functions+in+Nervous+System.">Eag1 K+ Channel: Endogenous Regulation and Functions in Nervous System.</a> Oxid. Med. Cell. Longev. 2017, (2017).</li><li>Han, B., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Human+EAG+channels+are+directly+modulated+by+PIP2+as+revealed+by+electrophysiological+and+optical+interference+investigations.">Human EAG channels are directly modulated by PIP2 as revealed by electrophysiological and optical interference investigations.</a> Sci. Rep. 6, (2016).</li><li>Do, T., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+rapid+method+for+determining+dynamic+binding+capacity+of+resins+for+the+purification+of+proteins.">A rapid method for determining dynamic binding capacity of resins for the purification of proteins.</a> PREP. 60 (2), 147-150 (2008).</li><li>Rohacs, T., Chen, J., Prestwich, G. D., Logothetis, D. E. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Distinct+specificities+of+inwardly+rectifying+K++channels+for+phosphoinositides.">Distinct specificities of inwardly rectifying K+ channels for phosphoinositides.</a> J. Biol. Chem. 274 (51), 36065-36072 (1999).</li><li>Overington, J. P., Al-Lazikani, B., Hopkins, A. L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=How+many+drug+targets+are+there?.">How many drug targets are there?.</a> Nat. Rev. Drug Discov. 5 (12), 993-996 (2006).</li><li>Suh, B. C., Hille, B. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=PIP2+is+a+necessary+cofactor+for+ion+channel+function:+How+and+why?.">PIP2 is a necessary cofactor for ion channel function: How and why?.</a> Annu. Rev. Biophys. 37, 175-195 (2008).</li><li>Yang, D., Singh, A., Wu, H., Kroe-Barrett, R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Determination+of+High-affinity+Antibody-antigen+Binding+Kinetics+Using+Four+Biosensor+Platforms.">Determination of High-affinity Antibody-antigen Binding Kinetics Using Four Biosensor Platforms.</a> J. Vis. Exp. (122), e55659 (2017).</li></ol>

膜离子通道已被证实为治疗各种人类疾病 (包括心血管和神经系统疾病) 的13% 以上的主要治疗靶点, 如18。膜片钳记录是测量小分子离子通道功能的黄金标准, 已广泛应用于离子通道配体的筛选。然而, 这种电生理方法无法证明小分子是否直接与通道绑定, 或者不是19, 因为小分子可以对与通道相互作用的其他蛋白质或细胞间通路采取行动。与广泛使用的放射性配体...

靶向细胞表面可接触的离子通道蛋白质与小分子提供了巨大的潜力, 配体筛选和生物药物发现1,2,3。因此, 研究离子通道与小分子之间的相互作用及其相应的作用是需要适当的工具。在离子通道功能检测中, 膜片钳记录已被证明是一种独特而不可替代的技术。然而, 确定小分子是否直接靶向离子通道需要其他技术。传统上, 采用放射性配体结合试验观察小分子与靶离子通道蛋白结合的动力学。然而, 由于其对放射性标记和检测的要求, 这种技术的使用受到限制。此外, 在研究中对小配体进行标记的先决条件是防止其在许多类型的离子通道中使用, 而不需要已知的特异配体。一些无标签的技术, 如核磁共振光谱学, x 射线衍射, 微型 thermophoresis (MST)4和表面等离子共振 (SPR) 已被用来测量蛋白质-小分子相互作用。但是这些类型的化验通常无法提供足够的信息, 因为很难获得全长蛋白质, 低分辨率的动力学, 低吞吐量和高成本的5。与这些技术相比, 生物层干涉法 (BLI) 正在成为一种新的无标签方法, 以克服这些缺陷, 通过固定少量蛋白质样品在表面上的蛋白质-小分子相互作用生物传感器和测量光学变化信号6,7。BLI 技术作为一种有前途的生物传感器平台, 已经进行了观察小分子与天然水溶性蛋白质 (如人单克隆抗体 CR80208 ) 的相互作用, 并报告了详细的化验程序。上一篇文章9。虽然离子通道蛋白在新的治疗靶点发现中的关键作用已经得到了确认, 但是基于 BLI 的离子通道蛋白-小分子相互作用的研究还没有被描述。
人类的以太hEAG1) 在各种类型的癌细胞和中枢神经系统中表达, 这使得该通道成为许多癌症和神经疾病的潜在治疗目标10,11, 12,13,14。我们实验室的电生理研究证实了磷酸肌醇4、5-bisphosphate (PIP2) 对 hEAG1通道15 的抑制作用。根据我们的结果, 测试 PIP2与 hEAG1 的直接交互使用 BLI 技术可以作为其他类型的离子通道蛋白-小分子化合物相互作用的模型, 特别是那些缺乏特定配体的通道。根据 BLI 化验的指示, 我们准备生物素化 hEAG1 蛋白, 并将它们固定在链亲和素 (SA) 生物传感器的表面上, 然后将它们与 PIP 2 解决方案相互作用, 观察它们之间的直接绑定。蛋白质和脂质。在将 PIP2附着到 hEAG1 蛋白涂层表面后, 表面层的厚度增加, 这直接关系到光谱的变化, 可以实时地测量16。由于关联步长的正移和离解步的负移位, 可以确定约束动力学。根据这一原理, 利用亲和纯化方法对 HEK-239T 稳定表达系统中的功能 hEAG1 离子通道蛋白进行纯化, 以维持体外功能状态, 然后测定其结合的动力学不同浓度 PIP2, 并产生了 semblable 的动力学数据, 如观察到的电生理测量15。BLI 和电生理测量结果之间的密切对应, 首次证明了 BLI 作为离子通道膜蛋白-小分子相互作用的适当分析工具的适用性。

<table>
	<tbody>
		<tr>
			<td>DMEM/High Glucose Medium</td>
			<td>HyClone</td>
			<td>SH30243.01</td>
			<td></td>
		</tr>
		<tr>
			<td>Phosphate Buffered Saline (1x)</td>
			<td>HyClone</td>
			<td>SH30256.01</td>
			<td></td>
		</tr>
		<tr>
			<td>Fetal Bovine Serum</td>
			<td>Gibco</td>
			<td>10270</td>
			<td></td>
		</tr>
		<tr>
			<td>Penicillin/Streptomycin</td>
			<td>Gibco</td>
			<td>1697550</td>
			<td></td>
		</tr>
		<tr>
			<td>Cell Culture Dish</td>
			<td>Corning</td>
			<td>430599</td>
			<td>150 mm X 25 mm</td>
		</tr>
		<tr>
			<td>Nonidet P-40 Substitute</td>
			<td>Amresco</td>
			<td>E109</td>
			<td></td>
		</tr>
		<tr>
			<td>Sodium chloride</td>
			<td>BBI Life Sciences</td>
			<td>A610476</td>
			<td></td>
		</tr>
		<tr>
			<td>Potassium chloride</td>
			<td>BBI Life Sciences</td>
			<td>A610440</td>
			<td></td>
		</tr>
		<tr>
			<td>Bovine Serum Albumin</td>
			<td>BBI Life Sciences</td>
			<td>A600332</td>
			<td></td>
		</tr>
		<tr>
			<td>Polyoxyethylene-20-Sorbitan Monolaurate</td>
			<td>BBI Life Sciences</td>
			<td>A600560</td>
			<td></td>
		</tr>
		<tr>
			<td>ANTI-FLAG M2 Affinity Gel</td>
			<td>Sigma</td>
			<td>A2220</td>
			<td></td>
		</tr>
		<tr>
			<td>3x FLAG peptide</td>
			<td>Sigma</td>
			<td>F4799</td>
			<td></td>
		</tr>
		<tr>
			<td>Octet-RED96</td>
			<td>Pall/Fort&#233;Bio</td>
			<td>30-5048</td>
			<td></td>
		</tr>
		<tr>
			<td>Data Acquisition software</td>
			<td>Pall/Fort&#233;Bio</td>
			<td>Version 7.1</td>
			<td></td>
		</tr>
		<tr>
			<td>Data Analysis software</td>
			<td>Pall/Fort&#233;Bio</td>
			<td>Version 7.1</td>
			<td></td>
		</tr>
		<tr>
			<td>Biosensor/Streptavidin</td>
			<td>Pall/Fort&#233;Bio</td>
			<td>18-5019</td>
			<td></td>
		</tr>
		<tr>
			<td>Microtiter plate</td>
			<td>Greiner Bio-one</td>
			<td>655209</td>
			<td></td>
		</tr>
		<tr>
			<td>Sulfo-NHS-LC-LC-Biotin</td>
			<td>ThermoFisher</td>
			<td>21338</td>
			<td></td>
		</tr>
		<tr>
			<td>Centrifugal Machine</td>
			<td>ThermoFisher</td>
			<td>75004250</td>
			<td></td>
		</tr>
		<tr>
			<td>PageRuler Prestained Protein Ladder</td>
			<td>ThermoScientific</td>
			<td>318120</td>
			<td></td>
		</tr>
		<tr>
			<td>Ultrafiltration device</td>
			<td>MILLIPORE</td>
			<td>UFC503008</td>
			<td>NMWL of 30 kDa</td>
		</tr>
		<tr>
			<td>phosphatidylinositol 4, 5-bisphosphate (PIP2)</td>
			<td>Sigma</td>
			<td>P9763</td>
			<td></td>
		</tr>
		<tr>
			<td>Monoclonal ANTI-FLAG M2 antibody</td>
			<td>Sigma</td>
			<td>F1804</td>
			<td>1:2000 dilution</td>
		</tr>
		<tr>
			<td>goat anti-mouse HRP-conjugated secondary antibody</td>
			<td>Santa Cruz Biotechnology</td>
			<td>sc-2005</td>
			<td>1:5000 dilution</td>
		</tr>
		<tr>
			<td>Enhanced BCA Protein Assay Kit</td>
			<td>Beyotime</td>
			<td>P0010</td>
			<td></td>
		</tr>
		<tr>
			<td>Protease Inhibitor Cocktail Tablets</td>
			<td>Roche</td>
			<td>04693159001</td>
			<td></td>
		</tr>
		<tr>
			<td>Amersham Imager 600 Imaging System</td>
			<td>GE Healthcare Bio-Sciences</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Western blot system</td>
			<td>BIO-RAD</td>
			<td></td>
			<td></td>
		</tr>
	</tbody>
</table>

capturing the interaction kinetics of an ion channel protein with small molecules by the bio-layer interferometry assay

生物层干涉法 (BLI) 是测定蛋白质和蛋白质-小分子相互作用的重要工具。在这里, 我们首先描述了这个新的无标签技术的应用研究人 EAG1 (hEAG1) 通道蛋白与小分子 PIP 的相互作用2。hEAG1 通道被认为是潜在的治疗靶点, 因为它在癌症中的异常过度表达, 以及某些类型的神经系统疾病所涉及的一些增益功能突变。我们从哺乳动物稳定表达系统中纯化 hEAG1 通道蛋白, 并用 BLI 测量与 PIP2的交互作用。hEAG1 蛋白与 PIP 的结合动力学的成功测量2表明 BLI 法是一种潜在的高通量方法, 用于离子通道药理学中新的小分子配体筛选。

我们从 HEK-293T 细胞稳定抗原 hEAG1 中纯化了旗融合 hEAG1 通道蛋白。采用膜片钳法对该融合蛋白的功能进行了验证, 纯化蛋白的质量和特异性由西方印迹 (图 1) 确定。纯化通道蛋白生物素化使用实时 BLI 检测与脂质 (PIP2) 进行交互试验。BLI 绑定检测配置显示在图 2中。hEAG1 和 PIP2之间的典型绑定曲线显示在<strong class...

Watch this Scientific Journal Video about Capturing the Interaction Kinetics of an Ion Channel Protein with Small Molecules by the Bio-layer Interferometry Assay at JoVE.com

Capturing the Interaction Kinetics of an Ion Channel Protein with Small Molecules by the Bio-layer Interferometry Assay

该协议描述了纯化 hEAG1 离子通道蛋白与小分子脂配体磷酸肌醇4、5-bisphosphate (PIP2) 的相互作用。实验表明, BLI 可能是一种新的小分子离子通道配体筛选的潜在方法。

用生物层干涉法捕获小分子离子通道蛋白的相互作用动力学

The bio-layer interferometry (BLI) assay is a valuable tool for measuring protein-protein and protein-small molecule interactions. Here, we first describe ...

Research

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Biochemistry

Method for Identifying Small Molecule Inhibitors of the Protein-protein Interaction Between HCN1 and TRIP8b

方法确定HCN1和TRIP8b之间蛋白 - 蛋白相互作用的小分子抑制剂

Hyperpolarization-activated cyclic nucleotide-gated (HCN) channels are expressed ubiquitously throughout the brain, where they function to regulate the ...

科研

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Visualization of Protein-protein Interaction in Nuclear and Cytoplasmic Fractions by Co-immunoprecipitation and In Situ Proximity Ligation Assay

Visualization of Protein-protein Interaction in Nuclear and Cytoplasmic Fractions by Co-immunoprecipitation and In Situ Proximity Ligation Assay

通过免疫共沉淀在核质馏分蛋白质相互作用的可视化和<em&gt;原位</em&gt;接近结扎分析

Protein-protein interactions are involved in thousands of cellular processes and occur in distinct spatial context. Traditionally, co-immunoprecipitation ...

Force Spectroscopy of Single Protein Molecules Using an Atomic Force Microscope

原子力显微镜对单蛋白质分子的力谱

The determination of the folding process of proteins from their amino acid sequence to their native 3D structure is an important problem in biology. ...

Measurement of Ion Concentration in the Unstirred Boundary Layer with Open Patch-Clamp Pipette: Implications in Control of Ion Channels by Fluid Flow

开放补丁夹具移液器测量未搅拌边界层中的离子浓度: 流体流动对离子通道控制的启示

Fluid flow is an important environmental stimulus that controls many physiological and pathological processes, such as fluid flow-induced vasodilation. ...

Ion Exchange Chromatography (IEX) Coupled to Multi-angle Light Scattering (MALS) for Protein Separation and Characterization

Ion Exchange Chromatography IEX Coupled to Multi-angle Light Scattering MALS for Protein Separation and Characterization

离子交换色谱 (IEX) 与多角度光散射 (ALS) 相结合, 用于蛋白质分离和表征

Ion-exchange chromatography with multi-angle light scattering (IEX-MALS) is a powerful method for protein separation and characterization. The combination ...

A Semi-High-Throughput Adaptation of the NADH-Coupled ATPase Assay for Screening Small Molecule Inhibitors

用于筛选小分子抑制剂的NADH耦合ATPase分析的半高通量自适应

ATPase enzymes utilize the free energy stored in adenosine triphosphate to catalyze a wide variety of endergonic biochemical processes in vivo that would ...

An In Vitro Single-Molecule Imaging Assay for the Analysis of Cap-Dependent Translation Kinetics

An In Vitro Single-Molecule Imaging Assay for the Analysis of Cap-Dependent Translation Kinetics

用于分析依赖帽的翻译动力学的 体外 单分子成像测定

Cap-dependent protein synthesis is the predominant translation pathway in eukaryotic cells. While various biochemical and genetic approaches have allowed ...

Derivatization of Protein Crystals with I3C using Random Microseed Matrix Screening

使用随机微种子基质筛选与 I3C 的蛋白质晶体的衍生

Protein structure elucidation using X-ray crystallography requires both high quality diffracting crystals and computational solution of the diffraction ...

Microcrystal Electron Diffraction of Small Molecules

小分子的微晶电子衍射

A detailed protocol for preparing small molecule samples for microcrystal electron diffraction (MicroED) experiments is described. MicroED has been ...

Identifying the Binding Proteins of Small Ligands with the Differential Radial Capillary Action of Ligand Assay (DRaCALA)

Identifying the Binding Proteins of Small Ligands with the Differential Radial Capillary Action of Ligand Assay DRaCALA

用利甘德分析（DRACALA）的微分径向毛细管作用识别小配体的结合蛋白

The past decade has seen tremendous progress in the understanding of small signaling molecules in bacterial physiology. In particular, the target proteins ...