Structural Biology of the Antitermination Process Regulating Transcription

Division Of Basic Sciences - Nci

Investigators

Abstract

Our studies have focused on the protein:protein and protein:RNA interactions associated with the NusB and NusE proteins and the boxA RNA, derived from binary complexes. We have determined the solution structure of NusB from Aquifex aeolicus, shown it to be consistent with all NusB homologues, and investigated the biophysics of NusB:NusE interactions. These studies have revealed the binding site of NusE on NusB and provide an explanation for the increased affinity for boxA RNA binding to the binary NusB:NusE complex compared to boxA RNA:NusB binding. This work was published in the Journal of Molecular Biology. The solution structure of the E. coli NusB:boxA RNA binary complex proved refractory;however we have been able to identify the binding site and orientation of the boxA RNA on NusB. Using a recently published crystal structure of NusB:NusE, this work reveals new insight into the ternary complex of NusB:NusE:boxA RNA. These studies are enhanced by analyses of the binding interactions for various known mutations of NusB. In parallel, we are pursuing direct structural determination of the ternary complex, utilizing stabilized orthologs and engineered proteins based on previous structural studies from our lab and others. In collaboration with the lab of Dr. Xinhua Ji, crystallographic studies of thermophilic homologues are providing further insight. The overall study will enable a more detailed and global picture of the interactions critical to the process of antitermination than have been available before. We plan to terminate this project following these structural studies.