HIGH PRESSURE COOLING OF E COLI CLASS IA RIBONUCLEOTIDE REDUCTASE COMPLEX

Cornell University, Ithaca NY

Investigators

Linked publications & trials

Abstract

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. Ribonucleotide reductases (RNR[unreadable][unreadable]"s) are the sole source of deoxyribonucletides in the cell an are thus vital for all living cells. While there are multiple classes of RNR[unreadable][unreadable]"s the E. coli class I enzyme has been extensively studied. The reaction requires two proteins, the active site containing alpha and the radical generating protein beta. Beta provides a radical to the active site of alpha, required for catalysis. While structures of the two subunits are known, there is no available structure of alpha and beta in complex and the oligomerization state of the active complex is not known. Crystals were obtained of alpha and beta in complex to begin to address some of the issues unresolved by biochemical and biophysical methods used so far. The best diffraction seen for these crystals is around 5.5 [unreadable] . A 5.8 [unreadable] data set was collected and the structure was solved at this resolution by molecular replacement using the two know subunit structures. The maps are of very high quality considering the low resolution. We are currently trying to improve the resolution in order to obtain atomic resolution information about the complex we do see in the structure. In collaboration with Dr. Sol M. Gruner and Dr. Chae Un Kim at Cornell, we are attempting to improve the resolution of our crystals using their newly developed high-pressure cryocooling approach. We are requesting beam time at CHESS to test the effects of high-pressure cryocooling, carried out at Cornell, on the diffraction properties of these crystals, and if successful, collecting full native datasets.