Name: analysis of athird11 intrinsic disorder and binding towards metal ions by capillary gel electrophoresis and affinity capillary electrophoresis
Uploaded: 2018-08-22T14:30:00
Description: Watch this Scientific Journal Video about Analysis of AtHIRD11 Intrinsic Disorder and Binding Towards Metal Ions by Capillary Gel Electrophoresis and Affinity Capillary Electrophoresis at JoVE.com

我们感谢 Masakuza (日本静冈县大学绿色科技研究院) 提供 AtHIRD11 蛋白质样品。

1. 毛细管电泳仪器的制备
<ol><li>
准备毛细血管
	<ol>
<li>使用玻璃切割机切割一个裸熔融二氧化硅毛细管与聚酰亚胺外涂层和内径为50µm 到33厘米 (为 CGE 实验) 和30厘米 (ACE 实验) 长毛细血管。在玻璃板上进行切割。 注: 为2种不同的仪器开发了2种不同的实验装置。为了在其他仪器上使用它们, 必须进行适当的方法转移, 并且必须将毛细管长度调整到仪器的允许长度。</li>
		<li>使用钢笔...

<ol>
	<li>Hara, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+multifunctionality+of+dehydrins:+An+overview.">The multifunctionality of dehydrins: An overview.</a> Plant Signaling &amp; Behavior. 5 (5), 1-6 (2010).</li><li>Uversky, V. N. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Dancing+protein+clouds:+the+strange+biology+and+chaotic+physics+of+intrinsically+disordered+proteins.">Dancing protein clouds: the strange biology and chaotic physics of intrinsically disordered proteins.</a> Journal of Biological Chemistry. 291 (13), 6681-6688 (2016).</li><li>Hara, M., 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=Biochemical+characterization+of+the+Arabidopsis+KS-type+dehydrin+protein,+whose+gene+expression+is+constitutively+abundant+rather+than+stress+dependent.">Biochemical characterization of the Arabidopsis KS-type dehydrin protein, whose gene expression is constitutively abundant rather than stress dependent.</a> Acta Physiologia Plantarum. 33 (6), 2103-2116 (2011).</li><li>Nachbar, M., 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=Metal+ion+-+dehydrin+interactions+investigated+by+affinity+capillary+electrophoresis+and+computer+models.">Metal ion - dehydrin interactions investigated by affinity capillary electrophoresis and computer models.</a> Journal of Plant Physiology. 216, 219-228 (2017).</li><li>Alhazmi, H. 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=A+comprehensive+platform+to+investigate+protein-metal+ion+interactions+by+affinity+capillary+electrophoresis.">A comprehensive platform to investigate protein-metal ion interactions by affinity capillary electrophoresis.</a> Journal of Pharmaceutical and Biomedical Analysis. 107, 311-317 (2015).</li><li>Redweik, S., Xu, Y., W&#228;tzig, H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Precise,+fast,+and+flexible+determination+of+protein+interactions+by+affinity+capillary+electrophoresis:+Part+1:+Performance.">Precise, fast, and flexible determination of protein interactions by affinity capillary electrophoresis: Part 1: Performance.</a> ELECTROPHORESIS. 33 (22), 3316-3322 (2012).</li><li>Ghosal, S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Electrokinetic+flow+and+dispersion+in+capillary+electrophoresis.">Electrokinetic flow and dispersion in capillary electrophoresis.</a> Annual Review of Fluid Mechanics. 38 (1), 309-338 (2006).</li><li>Xuan, X., Li, D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Analytical+study+of+Joule+heating+effects+on+electrokinetic+transportation+in+capillary+electrophoresis.">Analytical study of Joule heating effects on electrokinetic transportation in capillary electrophoresis.</a> Journal of Chromatography A. 1064 (2), 227-237 (2005).</li><li>Oledzka, I. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Comparative+evaluation+of+tissue+protein+separations+applying+one-+dimensional+gel+electrophoresis+and+capillary+gel+electrophoresis.">Comparative evaluation of tissue protein separations applying one- dimensional gel electrophoresis and capillary gel electrophoresis.</a> The Open Proteomics Journal. 5 (1), 17-21 (2012).</li><li>Alhazmi, H. 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=Optimization+of+affinity+capillary+electrophoresis+for+routine+investigations+of+protein-metal+ion+interactions.">Optimization of affinity capillary electrophoresis for routine investigations of protein-metal ion interactions.</a> Journal of Separation Science. 38 (20), 3629-3637 (2015).</li><li>Busch, M. H. A., Carels, L. B., Boelens, H. F. M., Kraak, J. C., Poppe, H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Comparison+of+five+methods+for+the+study+of+drug-protein+binding+in+affinity+capillary+electrophoresis.">Comparison of five methods for the study of drug-protein binding in affinity capillary electrophoresis.</a> Journal of Chromatography A. 777 (2), 311-328 (1997).</li><li>Mozafari, M., Balasupramaniam, S., Preu, L., El Deeb, S., Reiter, C. G., W&#228;tzig, H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Using+affinity+capillary+electrophoresis+and+computational+models+for+binding+studies+of+heparinoids+with+P-selectin+and+other+proteins.">Using affinity capillary electrophoresis and computational models for binding studies of heparinoids with P-selectin and other proteins.</a> ELECTROPHORESIS. 38 (12), 1560-1571 (2017).</li></ol>

对于所有的 CE 实验, 毛细管的制备是一个关键步骤。由于它是一个小直径的玻璃管, 它可以很容易地打破在涂层被删除时, 它正在处理。将毛细管安装到仪器中应该非常小心。
使用 ACE 方法所涉及的关键步骤主要与实验设置有关。另一个关键步骤是找到正确的样本注入参数。样品的注入量必须足够高的峰值。然而, 重载它将导致一个不太解决的峰值模式与重叠的峰值。在减小注...

植物比其他许多生命形式更依赖于他们的环境。由于植物不能迁移到其他地方, 它们必须适应周围环境的变化 (例如干旱、寒冷和高盐浓度)。因此, 高等植物开发了像 dehydrins 这样的特殊的应力蛋白, 它完成多种任务, 减少与高盐度有关的细胞应力。这些蛋白结合细胞内的水和离子, 通过结合铜2 +离子来减少氧化应激, 并与磷脂和骨架相互作用。此外, 结合锌2 +离子允许这些蛋白质作为转录因子。在磷酸化后, 它们结合钙2 +离子的能力也被报告了1。
这些蛋白质的多功能行为与无疏水性氨基酸残留有关。因此, 它们缺乏肽链内的任何疏水相互作用, 也没有受约束的结构。然而, 由于这些蛋白质缺乏限制性结构, 它们可以在相同的条件下占据不同的构象。因此, 它们可以被描述为结构的集合, 而不是单一的构造。这些性质的蛋白质被称为内在紊乱的蛋白质 (国内流离失所者), 是一个广泛使用的概念, 在应力蛋白和串扰之间的不同途径真核细胞在真核细胞2。
这些与压力有关的国内流离失所者之一是 AtHIRD11。它是拟南芥最干旱表达的国内流离失所者之一。因此, 不同的构象可以用其有效的半径与电荷比分开, 毛细管电泳 (CE) 已被用于进一步的研究。先前的 ACE 实验证明了 AtHIRD11 和过渡金属离子的相互作用, 如铜2 +-锌2 +-Co2 +和镍2 +离子。详细的结果可以在奥哈拉等地找到。3和 Nachbar等。4。
将在这里使用的 ACE 方法是基于我们以前发布的作品6。然而, 在蛋白质样品中加入 EOF 标记乙酰苯胺是不合适的。AtHIRD11 显示了广泛的峰值模式, 并将 EOF 标记添加到样本将伪装成两个峰值。因此, 标记在单独的运行中使用。在对约束行为进行检验之前, 证实了先前实验中发现的峰值来自不同的构象。因此, CGE 被用来区分蛋白质构象, 转化后的修饰蛋白, 和杂质, 如 AtHIRD11 的片段, 由其不同的质量。然后, 对不同金属离子的特征 AtHIRD11 样品的结合行为进行了研究。
本文的目的是描述一个实验设置, 以区分一个国内流离失所者和其他组成部分的样本, 以评估不同的绑定行为的构象。

<table border="0" cellpadding="0" cellspacing="0" width="638">
	<tbody>
		<tr height="84">
			<td height="84" style="height:84px;width:216px;">AtHIRD11 sample</td>
			<td style="width:111px;">Shizuoka University (Group Prof. M. Hara)</td>
			<td style="width:119px;">-</td>
			<td style="width:192px;">Dehydrin Protein from Arabidopsis thaliana expressed in Escherichia coli</td>
		</tr>
		<tr height="63">
			<td height="63" style="height:63px;width:216px;">Barefused silica capillary</td>
			<td style="width:111px;">Polymicro Technologies (Phoenix, USA)</td>
			<td style="width:119px;">106815-0017</td>
			<td style="width:192px;">TSP050375, 50 &#956;m inner diameter, 363 &#956;m outer diameter, polyimide coating</td>
		</tr>
		<tr height="84">
			<td height="84" style="height:84px;width:216px;">Agilent 1600A</td>
			<td style="width:111px;">Agilent Technologies (Waldbronn, Germany)</td>
			<td style="width:119px;">comercially not available anymore</td>
			<td style="width:192px;">Capillary electrophoresis instrument; Agilent 7100 CE can be used instead</td>
		</tr>
		<tr height="84">
			<td height="84" style="height:84px;width:216px;">Agilent 7100 CE</td>
			<td style="width:111px;">Agilent Technologies (Waldbronn, Germany)</td>
			<td style="width:119px;">G7100A</td>
			<td style="width:192px;">Capillary electrophoresis instrument</td>
		</tr>
		<tr height="63">
			<td height="63" style="height:63px;width:216px;">Injekt 2 mL</td>
			<td style="width:111px;">B. Braun (Melsungen, Germany)</td>
			<td style="width:119px;">4606051V</td>
			<td style="width:192px;">Syringe for filtration</td>
		</tr>
		<tr height="84">
			<td height="84" style="height:84px;width:216px;">Rotilabo-syringe filters</td>
			<td style="width:111px;">Carl Roth GmbH + Co. KG (Karlsruhe, Germany)</td>
			<td style="width:119px;">KY62.1</td>
			<td style="width:192px;">PVDF membrane filter for solution filtration</td>
		</tr>
		<tr height="84">
			<td height="84" style="height:84px;width:216px;">Eppendorf Research plus 10 &#956;L</td>
			<td style="width:111px;">Eppendorf (Wesseling-Berzdorf, Germany)</td>
			<td style="width:119px;">3121 000.023</td>
			<td style="width:192px;">Micro pipette for sample handling</td>
		</tr>
		<tr height="84">
			<td height="84" style="height:84px;width:216px;">Eppendorf Research plus 10 &#956;L</td>
			<td style="width:111px;">Eppendorf (Wesseling-Berzdorf, Germany)</td>
			<td style="width:119px;">3121 000.120</td>
			<td style="width:192px;">Micro pipette for handling the ligand solutions</td>
		</tr>
		<tr height="63">
			<td height="63" style="height:63px;width:216px;">Bulb pipette 10 mL</td>
			<td style="width:111px;">Duran Group GmbH(Mainz, Germany)</td>
			<td style="width:119px;">24 338 08</td>
			<td style="width:192px;">Preparing theNaOH solution</td>
		</tr>
		<tr height="63">
			<td height="63" style="height:63px;width:216px;">Bulb pipette 25 mL</td>
			<td style="width:111px;">Duran Group GmbH(Mainz, Germany)</td>
			<td style="width:119px;">24 338 14</td>
			<td style="width:192px;">Preparing the ligand solution</td>
		</tr>
		<tr height="63">
			<td height="63" style="height:63px;width:216px;">Duran glas volumetric flask 25 mL</td>
			<td style="width:111px;">Duran Group GmbH(Mainz, Germany)</td>
			<td style="width:119px;">24 671 1457</td>
			<td style="width:192px;">Preparing the ligand stock solution</td>
		</tr>
		<tr height="63">
			<td height="63" style="height:63px;width:216px;">Duran glas volumetric flask 10 mL</td>
			<td style="width:111px;">Duran Group GmbH(Mainz, Germany)</td>
			<td style="width:119px;">24 671 1054</td>
			<td style="width:192px;">Preparing the ligand stock solution</td>
		</tr>
		<tr height="147">
			<td height="147" style="height:147px;width:216px;">Proteome Lab SDS MW Gel Buffer</td>
			<td style="width:111px;">Beckman Coulter (Brea, USA)</td>
			<td style="width:119px;">comercially not available anymore</td>
			<td style="width:192px;">Separation during capillary gel electrophoresis / Alternative SDS buffer: CE-SDS run buffer from Bio-Rad Laboratories (M&#252;nchen, Germany) Catalog Number: 1485032&#160;</td>
		</tr>
		<tr height="63">
			<td height="63" style="height:63px;width:216px;">Acetanilide</td>
			<td style="width:111px;">Sigma-Aldrich (Steinheim, Germany)</td>
			<td style="width:119px;">397229-5G</td>
			<td style="width:192px;">Electroosmotic flow marker</td>
		</tr>
		<tr height="63">
			<td height="63" style="height:63px;width:216px;">Manganese(II) chloride</td>
			<td style="width:111px;">Sigma-Aldrich (Steinheim, Germany)</td>
			<td style="width:119px;">13217</td>
			<td style="width:192px;">Ligand</td>
		</tr>
		<tr height="63">
			<td height="63" style="height:63px;width:216px;">Ethylenediaminetetraacetic acid (EDTA)</td>
			<td style="width:111px;">Sigma-Aldrich (Steinheim, Germany)</td>
			<td style="width:119px;">431788-100G</td>
			<td style="width:192px;">Rinsing ingredient</td>
		</tr>
		<tr height="63">
			<td height="63" style="height:63px;width:216px;">Bariumchloride</td>
			<td style="width:111px;">Sigma-Aldrich (Steinheim, Germany)</td>
			<td style="width:119px;">202738-5G</td>
			<td style="width:192px;">Ligand</td>
		</tr>
		<tr height="63">
			<td height="63" style="height:63px;width:216px;">Sodium dodecyl sulfate</td>
			<td style="width:111px;">Sigma-Aldrich (Steinheim, Germany)</td>
			<td style="width:119px;">71729-100G</td>
			<td style="width:192px;">Solublizing protein for capillary gel electrophoresis</td>
		</tr>
		<tr height="63">
			<td height="63" style="height:63px;width:216px;">Nickel(II) chloride hexahydrate</td>
			<td style="width:111px;">Sigma-Aldrich (Steinheim, Germany)</td>
			<td style="width:119px;">654507-5G</td>
			<td style="width:192px;">Ligand</td>
		</tr>
		<tr height="63">
			<td height="63" style="height:63px;width:216px;">Selenium(IV) chloride</td>
			<td style="width:111px;">Sigma-Aldrich (Steinheim, Germany)</td>
			<td style="width:119px;">323527-10G</td>
			<td style="width:192px;">Ligand</td>
		</tr>
		<tr height="63">
			<td height="63" style="height:63px;width:216px;">2-amino-2-hydroxy-methylpropane-1.3-diol (Tris)</td>
			<td style="width:111px;">Sigma-Aldrich (Steinheim, Germany)</td>
			<td style="width:119px;">252859-100G</td>
			<td style="width:192px;">Buffer ingredient</td>
		</tr>
		<tr height="63">
			<td height="63" style="height:63px;width:216px;">Zinc(II) chloride</td>
			<td style="width:111px;">Merck Millipore ( Darmstadt, Germany)</td>
			<td style="width:119px;">1088160250</td>
			<td style="width:192px;">Ligand</td>
		</tr>
		<tr height="63">
			<td height="63" style="height:63px;width:216px;">Strontium nitrate</td>
			<td style="width:111px;">Merck Millipore ( Darmstadt, Germany)</td>
			<td style="width:119px;">1078720250</td>
			<td style="width:192px;">Ligand</td>
		</tr>
		<tr height="63">
			<td height="63" style="height:63px;width:216px;">Calcium chloride dihydrate</td>
			<td style="width:111px;">Merck Millipore ( Darmstadt, Germany)</td>
			<td style="width:119px;">1371015000</td>
			<td style="width:192px;">Ligand</td>
		</tr>
		<tr height="63">
			<td height="63" style="height:63px;width:216px;">37% hydrochloric acid</td>
			<td style="width:111px;">Merck Millipore ( Darmstadt, Germany)</td>
			<td style="width:119px;">1003171000</td>
			<td style="width:192px;">Adjusting pH</td>
		</tr>
		<tr height="63">
			<td height="63" style="height:63px;width:216px;">Copper(II) chloride dihydrate</td>
			<td style="width:111px;">Riedel-de Ha&#235;n (Seelze, Germany)</td>
			<td style="width:119px;">31286</td>
			<td style="width:192px;">Ligand</td>
		</tr>
		<tr height="63">
			<td height="63" style="height:63px;width:216px;">Sonorex Longlife RK 1028 CH 45L</td>
			<td style="width:111px;">Allpax (Papenburg, Germany)</td>
			<td style="width:119px;">10000084;0</td>
			<td style="width:192px;">Ultrasonic bath</td>
		</tr>
		<tr height="84">
			<td height="84" style="height:84px;width:216px;">Agilent ChemStation Rev. 8.04.03-SP1</td>
			<td style="width:111px;">Agilent Technologies (Waldbronn, Germany)</td>
			<td>G2070-91126</td>
			<td style="width:192px;">Software packages to operate the CE instruments, acquisite data and evaluate it</td>
		</tr>
	</tbody>
</table>

analysis of athird11 intrinsic disorder and binding towards metal ions by capillary gel electrophoresis and affinity capillary electrophoresis

植物强烈依赖于他们的环境。为了适应压力变化 (例如干旱和高盐度), 高等植物进化出属于内在紊乱的蛋白质 (国内流离失所者), 以减少氧化和渗透胁迫。本文采用毛细管凝胶电泳 (CGE) 和移动性移位亲和电泳 (ACE) 相结合的方式来描述拟南芥不同构象的 AtHIRD11 的结合行为。CGE 用于确认 AtHIRD11 的纯度, 并排除片段、翻译后修饰和其他杂质作为复杂峰值模式的原因。在这一部分的实验中, 不同的样品成分是由一个粘性凝胶在毛细管内的不同质量和检测与二极管阵列探测器。然后, 用 ACE 研究了样品对各种金属离子的结合行为。在这种情况下, 配体被添加到缓冲区解决方案中, 并测量迁移时间的偏移量, 以确定是否发生了绑定事件。使用 CGE 和 ACE 的组合来确定国内流离失所者的结合行为的优点之一是, 有可能自动进行凝胶电泳和结合试验。此外, 专家咨询小组显示的检测限制比经典凝胶电泳和 ACE 能够确定的方式, 结合一个配体的快速方式。此外, ACE 也可以应用于其他带电的物种比金属离子。但是, 使用这种方法进行绑定实验, 其确定绑定站点数量的能力受到限制。然而, 专家咨询小组和 ACE 的结合可以适应于描述任何蛋白质样品对许多带电配体的结合行为。

图 1显示了在 CGE 实验中获得的 AtHIRD11 样品的电泳图谱。肽的大小从左向右增加。峰值4有最大的质量和表明完整的蛋白质。较小的峰值2和3代表较小的杂质 (如降解产物)。在没有蛋白质样本的情况下, 第一峰值和基线的不一致也可以复制。因此, 它与 SDS 凝胶本身有关, 不代表任何与样品相关的杂质。
<p class="jove_content" fo:keep-together.within-page="1...

Watch this Scientific Journal Video about Analysis of AtHIRD11 Intrinsic Disorder and Binding Towards Metal Ions by Capillary Gel Electrophoresis and Affinity Capillary Electrophoresis at JoVE.com

Analysis of AtHIRD11 Intrinsic Disorder and Binding Towards Metal Ions by Capillary Gel Electrophoresis and Affinity Capillary Electrophoresis

该协议结合毛细管电泳对蛋白质样品的特性进行了表征, 并对带电配体进行了快速结合筛选。推荐用于具有弹性结构的蛋白质, 如内在紊乱的蛋白质, 以确定不同构象的结合的差异。

毛细管电泳与亲和毛细管电泳对金属离子 AtHIRD11 固有紊乱与结合的分析

Plants are strongly dependent on their environment. In order to adjust to stressful changes (e.g., drought and high salinity), higher plants evolve ...

Research

JoVE Journal

Biochemistry

Sheathless Capillary Electrophoresis&#8211;Mass Spectrometry for Metabolic Profiling of Biological Samples

Sheathless Capillary ElectrophoresisÃƒÆ’Ã‚Â¢ÃƒÂ¢Ã¢â‚¬Å¡Ã‚Â¬ÃƒÂ¢Ã¢â€šÂ¬Ã…â€œMass Spectrometry for Metabolic Profiling of Biological Samples

鞘毛细管电泳 - 质谱联用生物样品的代谢谱

In metabolomics, a wide range of analytical techniques is used for the global profiling of (endogenous) metabolites in complex samples. In this paper, a ...

科研

生物化学

Characterization of Multi-subunit Protein Complexes of Human MxA Using Non-denaturing Polyacrylamide Gel-electrophoresis

使用非变性聚丙烯酰胺凝胶电泳人的MxA多亚基蛋白复合物的表征

The formation of oligomeric complexes is a crucial prerequisite for the proper structure and function of many proteins. The interferon-induced antiviral ...

Capillary Electrophoresis Separation of Monoclonal Antibody Isoforms Using a Neutral Capillary

使用中性毛细管单克隆抗体亚型的毛细管电泳分离

Biotherapeutic proteins, such as monoclonal antibodies (mAbs), are feasible alternatives for the treatment of chronic-degenerative diseases. The ...

Determination of High-affinity Antibody-antigen Binding Kinetics Using Four Biosensor Platforms

高亲和力抗体 - 抗原的结合动力学的测定，使用四个生物传感器平台

Label-free optical biosensors are powerful tools in drug discovery for the characterization of biomolecular interactions. In this study, we describe the ...

Analyzing Supercomplexes of the Mitochondrial Electron Transport Chain with Native Electrophoresis, In-gel Assays, and Electroelution

用天然电泳，凝胶电泳和电解法分析线粒体电子传递链的超复合物

The mitochondrial electron transport chain (ETC) transduces the energy derived from the breakdown of various fuels into the bioenergetic currency of the ...

Horizontal Gel Electrophoresis for Enhanced Detection of Protein-RNA Complexes

水平凝胶电泳用于增强蛋白质 -  RNA复合物的检测

Native polyacrylamide gel electrophoresis is a fundamental tool of molecular biology that has been used extensively for the biochemical analysis of ...

DNA Polymerase Activity Assay Using Near-infrared Fluorescent Labeled DNA Visualized by Acrylamide Gel Electrophoresis

用丙烯酰胺凝胶电泳法可视化近红外荧光标记 dna 聚合酶活性

For any enzyme, robust, quantitative methods are required for characterization of both native and engineered enzymes. For DNA polymerases, DNA synthesis ...

Highly Sensitive and Quantitative Detection of Proteins and Their Isoforms by Capillary Isoelectric Focusing Method

毛细管电聚焦法对蛋白质及其亚型的高灵敏度和定量检测

Immunoblotting has become a routine technique in many laboratories for protein characterization from biological samples. The following protocol provides ...

Use of Two Dimensional Semi-denaturing Detergent Agarose Gel Electrophoresis to Confirm Size Heterogeneity of Amyloid or Amyloid-like Fibers

二维半变性洗涤剂琼脂糖凝胶电泳法确定淀粉样或淀粉样纤维的粒度异质性

Amyloid or amyloid-like fibers have been associated with many human diseases, and are now being discovered to be important for many signaling pathways. ...

Analysis of Thylakoid Membrane Protein Complexes by Blue Native Gel Electrophoresis

蓝色本机凝胶电泳法分析囊体膜蛋白配合物

Photosynthetic electron transfer chain (ETC) converts solar energy to chemical energy in the form of NADPH and ATP. Four large protein complexes embedded ...