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Modularity of Multiprotein Complexes and Protein Complex Architectures

We have begun to utilize deletion network analyses to propose the architecture of multiprotein complexes and to determine the modularity of protein complexes. In our initial publication in this area, we analyzed the Rpd3/Sin3 histone deacetylase complexes. Here, Rpd3 was used as an affinity purification bait and protein complexes were analyzed from a wild type background and in strains where different subunits of the complex where deleted. Using this approach we were able to propose an architectural model for the Rpd3/Sin3 histone deacetylase complexes (Sardiu et al., 2009). In this study, all results were obtained using a single TAP tagged bait, therefore the deletions can only be interpreted relative to the protein that was TAP tagged. This only provides insights into the local architecture of the complex around the TAP tagged protein and not for the whole complex. As a result, in certain deletions where the majority of the complex proteins were lost after deletion of the subunit, this approach was not able to distinguish if the deletion simply prevented the bait protein from binding to an otherwise intact complex or if the whole complex fell apart.

To overcome these limitations, we have improved upon the method with a study of the SAGA and ADA chromatin remodeling complexes (Lee et al., 2011). Here we used multiple bait protein purifications in combination with different deletions. With this combinatorial approach were able to actually purify and analyze sub modules of these protein complexes. In addition, we were able to determine which proteins are critical for complete complex assembly. From this dataset, 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.

Lee, K.K., Sardiu, M.E., Swanson, S.K., Gilmore, J.M., Torol, M., Grant, P.A., Grant, P., Florens, L., Workman, J.L. & Washburn, M.P. (2011) Combinatorial Depletion Analysis to Assemble the Network Architecture of the SAGA and ADA Chromatin Remodeling Complexes. Mol. Syst. Biol., 7:503. Abstract

Mosley, A.L., Sardiu, M.E., Pattenden, S.G., Workman, J.L., Florens, L., & Washburn, M.P. (2011) Highly reproducible label free quantitative proteomic analysis of RNA polymerase complexes. Mol. Cell. Proteomics, 10(2):M110.000687. Abstract

Sardiu, M.E., Gilmore, J.M., Carrozza, M.J., Li, B., Workman, J.L., Florens, L., & Washburn, M.P. (2009) Determining protein complex connectivity using a probabilistic deletion network derived from quantitative proteomics. PloS ONE, 4(10):e7310. Abstract