Microscale thermophoresis quantifies biomolecular interactions under previously challenging conditions.
Microscale thermophoresis quantifies biomolecular interactions under previously challenging conditions.
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DOI：
10.1016/j.ymeth.2012.12.005
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发表时间：
2013-03
期刊：
影响因子：
4.8
通讯作者：
Duhr, Stefan
中科院分区：
文献类型：
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作者：
Seidel, Susanne A. I.;Dijkman, Patricia M.;Lea, Wendy A.;van den Bogaart, Geert;Jerabek-Willemsen, Moran;Lazic, Ana;Joseph, Jeremiah S.;Srinivasan, Prakash;Baaske, Philipp;Simeonov, Anton;Katritch, Ilia;Melo, Fernando A.;Ladbury, John E.;Schreiber, Gideon;Watts, Anthony;Braun, Dieter;Duhr, Stefan
微量热泳（MST）允许定量分析蛋白质相互作用的自由溶液中，并与低样品消耗。该技术是基于热泳，定向运动的分子在温度梯度。热泳是高度敏感的所有类型的结合诱导的分子性质的变化，无论是在大小，电荷，水化壳或构象。在全光学方法中，使用红外激光进行局部加热，并通过荧光分析温度梯度中的分子迁移率。在标准MST中，一个结合配偶体被荧光标记。然而，MST也可以通过利用内在蛋白质UV-荧光进行无标记。尽管分子量比很高，但小分子和肽与蛋白质的相互作用很容易通过MST获得。此外，MST测定具有高度适应性，以适应不同生物分子的不同要求，例如待在溶液中稳定的膜蛋白。缓冲剂和添加剂的类型可以自由选择。甚至可以在复杂的生物液体（如细胞裂解物）中进行测量，从而接近体内条件，无需样品纯化。可通过MST定量的结合模式包括二聚化、协同性和竞争性。因此，它的灵活性，在试验设计资格MST的生物分子相互作用的分析在复杂的实验环境中，我们在此证明，解决典型的挑战性类型的结合事件，从生命科学的各个领域。
Microscale thermophoresis (MST) allows for quantitative analysis of protein interactions in free solution and with low sample consumption. The technique is based on thermophoresis, the directed motion of molecules in temperature gradients. Thermophoresis is highly sensitive to all types of binding-induced changes of molecular properties, be it in size, charge, hydration shell or conformation. In an all-optical approach, an infrared laser is used for local heating, and molecule mobility in the temperature gradient is analyzed via fluorescence. In standard MST one binding partner is fluorescently labeled. However, MST can also be performed label-free by exploiting intrinsic protein UV-fluorescence. Despite the high molecular weight ratio, the interaction of small molecules and peptides with proteins is readily accessible by MST. Furthermore, MST assays are highly adaptable to fit to the diverse requirements of different biomolecules, such as membrane proteins to be stabilized in solution. The type of buffer and additives can be chosen freely. Measuring is even possible in complex bioliquids like cell lysate allowing close to in vivo conditions without sample purification. Binding modes that are quantifiable via MST include dimerization, cooperativity and competition. Thus, its flexibility in assay design qualifies MST for analysis of biomolecular interactions in complex experimental settings, which we herein demonstrate by addressing typically challenging types of binding events from various fields of life science.
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影响因子：
5.7
作者：
Kufareva, Irina;Rueda, Manuel;Katritch, Vsevolod;Stevens, Raymond C.;Abagyan, Ruben;Yoshikawa, Yasushi;Furuya, Toshio;Lee, Huisun;Roy, Ambrish;Grime, John;Rebehmed, Joseph;Zhang, Yang;Roumen, Luc;de Esch, Iwan J. P.;Leurs, Rob;de Graaf, Chris;Li, Youyong;Hou, Tingjun;Mysinger, Michael M.;Weiss, Dahlia R.;Irwin, John J.;Shoichet, Brian K.;McRobb, Fiona M.;Capuano, Ben;Crosby, Ian T.;Chalmers, David K.;Yuriev, Elizabeth;Wang, Qi;Mach, Robert H.;Reichert, David E.;Chuang, Gwo-Yu;Rognan, Didier;Simms, John;Sexton, Patrick;Wootten, Denise;Latek, Dorota;Ghoshdastider, Umesh;Filipek, Slawomir;Kirkpatrick, Andrea;Trzaskowski, Bartosz;Griffith, Adam;Kim, Soo-Kyung;Abrol, Ravinder;Goddard, William A., III;Vaidehi, Nagarajan;Lam, Alfonso;Bhattacharya, Supriyo;Li, Hubert;Balaraman, Gouthaman;Niesen, Michiel;Pal, Sandeep;Vorobjev, Yrii;Bakulina, Natalia;Solovyev, Victor;Beuming, Thijs;Costanzi, Stefano;Shi, Lei;Higgs, Chris;Salam, Noeris;Lupyan, Dmitry;Sherman, Woody;Ding, Feng;Kota, Pradeep;Ramachandran, Srinivas;Dokholyan, Nikolay V.;Carlsson, Jens;Coleman, Ryan G.;Fan, Hao;Schlessinger, Avner;Sali, Andrej;Tikhonova, Irina;Pogozheva, Irina;Lomize, Andrei;Hall, Nathan E.;Muddassar, Muhammad;Zhang, Yang;Pae, Ae Nim;Lee, Jooyoung;Lopez, Laura;Obiol-Pardo, Cristian;Selent, Jana;Mahboob, Sadia;Werner, Tim;Church, W. Bret;Brylinski, Michal;Ando, Tadashi;Guerler, Aysam;Zhou, Hongyi;Skolnick, Jeffrey;Xhaard, Henri;Jurkowski, Wiktor;Elofsson, Arne;Murad, Ahsan K.;Drwal, Malgorzata;Dupree, Tom B.;Griffith, Renate;Ostopovici-Halip, Liliana;Bologa, Cristian;Chen, K. M.;Sun, J.;Barth, Patrick;Yarov-Yarovoy, Vladimir;Baker, David;Vroling, Bas;Sanders, Marijn P. A.;Nabuurs, Sander B.;Nikiforovich, Gregory V.
通讯作者：
Nikiforovich, Gregory V.
DOI：
10.1073/pnas.0603873103
发表时间：
2006-12-26
影响因子：
11.1
作者：
Duhr, Stefan;Braun, Dieter
通讯作者：
Braun, Dieter
影响因子：
3.6
作者：
Fredriksson, R;Schiöth, HB
通讯作者：
Schiöth, HB
影响因子：
16
作者：
Kubicek, Stefan;O'Sullivan, Roderick J.;Jenuwein, Thomas
通讯作者：
Jenuwein, Thomas
影响因子：
46.9
作者：
通讯作者：
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