NSF Award
9506933
9506933 Assa-Munt Transcriptional regulation is modulated by the interaction between transcription factors and their target DNA sequences, as well as by protein protein interactions. Escherichia coli RNA polymerase holoenzyme (RNAP) is one of the most intensely studied enzymes since its discovery. This multimeric enzyme consists of a core protein made up of 2 ' and complexes with the major sigma subunit (sigma 70). This research will concentrate on the role and structure of the subunit of RNA polymerase, and specifically on the carboxyterminal domain of , ( -CTD). This domain is known to bind DNA and also to interact with CAP, the catabolite gene activator protein. Its small size makes it amenable to NMR solution studies and it has been expressed in large amounts. The solution structure of the -CTD will be determined by multidimensional NMR studies, both free in solution and when complexed to DNA. Dynamic studies will probe the role of flexible regions in the protein and also in the protein-DNA complex. Based on the solved structure, we will identify the topological surfaces involved in protein-protein and in protein-DNA recognition, and use the structures to understand the available biochemical and mutation data. The -CTD is an independently folded domain and may be studied as a prototype of the dual DNA-binding and protein-protein functions common to both eukaryotic and prokaryotic DNA-binding proteins. %%% Transcriptional regulation depends on the interaction between transcription factors and their target DNA sequences. Modulation of the biological activity of these transcription factors via protein-DNA or protein-protein interactions regulates processes such as cell development, differentiation and oncogenesis. One of the best studied transcription systems is the bacterial RNA polymerase and its interactions with diverse promoters. This research on fundamental issues of DNA-binding proteins and their regulatory roles mediated by specific pro tein-protein and protein- DNA associations will focus on the domain organization of the RNA- polymerase subunit from E. coli; it binds DNA and interacts with the activating region of the Catabolite Gene Activator protein (CAP) in the ternary complex of the lac promoter, RNAP and CAP. It is, therefore, a good model to study intermolecular interactions. ***
