Combinatorial depletion analysis to assemble the network architecture of the SAGA and ADA chromatin remodeling complexes.
Combinatorial depletion analysis to assemble the network architecture of the SAGA and ADA chromatin remodeling complexes.
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DOI：
10.1038/msb.2011.40
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发表时间：
2011-07-05
影响因子：
9.9
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中科院分区：
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一个组合耗尽策略与生物化学，定量蛋白质组学和计算方法相结合，以阐明SAGA/ADA复合物的结构。分析揭示了五个连接的功能模块能够独立组装。尽管有几个大规模的蛋白质组学研究，旨在确定全球范围内的蛋白质相互作用的可用性，很少有人知道蛋白质如何相互作用，并在大分子复合物内组织。在这里，我们描述了一种技术，包括生物化学方法，定量蛋白质组学和计算方法的组合，使用野生型和缺失菌株，以调查大分子蛋白质复合物内的蛋白质的组织。我们应用这种技术来确定组织的两个充分研究的复合物，Spt-Ada-Gcn 5组蛋白乙酰转移酶（SAGA）和ADA，其中没有全面的高分辨率结构存在。该方法揭示了SAGA/ADA由五个不同的功能模块组成，它们可以单独持久化。此外，我们确定了一个新的亚基的ADA复合物，称为Ahc 2，并表征Sgf29作为ADA家族蛋白存在于所有Gcn 5组蛋白乙酰转移酶复合物。最后，我们提出了一个模型的架构的SAGA和ADA复合物，预测新的功能协会内的SAGA复合物，并提供了机械的见解，在SAGA突变体的表型观察。
A combinatorial depletion strategy is combined with biochemistry, quantitative proteomics and computational approaches to elucidate the structure of the SAGA/ADA complexes. The analysis reveals five connected functional modules capable of independent assembly. Despite the availability of several large-scale proteomics studies aiming to identify protein interactions on a global scale, little is known about how proteins interact and are organized within macromolecular complexes. Here, we describe a technique that consists of a combination of biochemistry approaches, quantitative proteomics and computational methods using wild-type and deletion strains to investigate the organization of proteins within macromolecular protein complexes. We applied this technique to determine the organization of two well-studied complexes, Spt–Ada–Gcn5 histone acetyltransferase (SAGA) and ADA, for which no comprehensive high-resolution structures exist. This approach revealed that SAGA/ADA is composed of five distinct functional modules, which can persist separately. Furthermore, we identified a novel subunit of the ADA complex, termed Ahc2, and characterized Sgf29 as an ADA family protein present in all Gcn5 histone acetyltransferase complexes. Finally, we propose a model for the architecture of the SAGA and ADA complexes, which predicts novel functional associations within the SAGA complex and provides mechanistic insights into phenotypical observations in SAGA mutants.