Analysis of stable and transient protein-protein interactions.
Analysis of stable and transient protein-protein interactions.
复制标题
DOI：
10.1007/978-1-61779-477-3_10
复制
发表时间：
2012
期刊：
影响因子：
--
通讯作者：
Tackett, Alan J
中科院分区：
文献类型：
--
作者：
Byrum, Stephanie;Smart, Sherri K;Larson, Signe;Tackett, Alan J
蛋白质组装成确定的复合物驱动了大量的细胞活动。这些蛋白质复合物通常具有一组更稳定的相互作用蛋白质以及更不稳定或瞬时的相互作用。研究这些蛋白质复合物的体内组分是具有挑战性的，因为许多用于分离的技术导致仅纯化最稳定的组分，并且瞬时相互作用丢失。一种称为瞬时同位素区分相互作用为随机或靶向的技术（瞬时I-DIRT）已被开发用于识别这些瞬时相互作用的蛋白质以及稳定的相互作用。这里描述的是用于多亚基复合物，NuA 3，乙酰化组蛋白H3和功能激活基因转录的瞬时I-DIRT分析的详细方法学途径。已知转录涉及在染色质/基因模板上执行不同活性的蛋白质组装的音乐会，因此理解与NuA 3的不太稳定或瞬时的蛋白质相互作用将揭示协同或拮抗地发挥作用以调节基因转录和染色质重塑的蛋白质复合物。
The assembly of proteins into defined complexes drives a plethora of cellular activities. These protein complexes often have a set of more stably interacting proteins as well as more unstable or transient interactions. Studying the in vivo components of these protein complexes is challenging as many of the techniques used for isolation result in the purification of only the most stable components and the transient interactions are lost. A technology called transient isotopic differentiation of interactions as random or targeted (transient I-DIRT) has been developed to identify these transiently interacting proteins as well as the stable interactions. Described here are the detailed methodological approaches used for a transient I-DIRT analysis of a multi-subunit complex, NuA3, that acetylates histone H3 and functions to activate gene transcription. Transcription is known to involve a concert of protein assemblies performing different activities on the chromatin/gene template, thus understanding the less stable or transient protein interactions with NuA3 will shed light onto the protein complexes that function synergistically, or antagonistically, to regulate gene transcription and chromatin remodeling.