This subproject is one of many research subprojects utilizing theresources provided by a Center grant funded by NIH/NCRR. The subproject andinvestigator (PI) may have received primary funding from another NIH source,and thus could be represented in other CRISP entries. The institution listed isfor the Center, which is not necessarily the institution for the investigator.Mediator is a central regulatory complex in transcription, and transmits information from activators and repressors bound at the regulatory DNA elements to RNA polymerase II (Pol II) in the entire range of organisms from yeast to humans. Mediator is composed of a total of 21 subunits with a molecular mass of over a million Dalton. We have successfully co-expressed all seven subunits of the head module of Mediator (223 kDa), and purified it in a large quantity. We have recently obtained crystals diffracting up to 4 , and a full native data set at 5.6 . We turned to small angle x-ray scattering to obtain an initial phase model for the crystal structure determination as well as to study the full-length Mediator, its subcomponents such as the head module, as well as the Mediator-Pol II complex. We have recently recorded solution x-ray scattering data to test feasibility of obtaining a structure at intermediary resolution. The Mediator head module sample in an optimal condition suffered little radiation damage. We obtained the radius of gyration 52 , the maximum particle dimension of 165 . We have been able to obtain a three dimensional model using Dammin and data recorded up to Q~0.26 -1. Remarkably, the shape of the solution structure is consistent with the structure of the head module of Mediator determined by electron microscopy and single particle analysis. More recently, we obtained beam time on our rapid access proposal, and extended the structural resolution to approximately 10 . One of the structural envelopes obtained by Gasbor using the most recent data has given an encouraging result in the crystallographic phase extension using Phaser.
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