The growth, development, and metabolism of healthy cells depends on the precise regulation of gene expression and error-free maintenance of the genome. A detailed understanding of the underlying principles of protein-DNA recognition and protein-DNA architectures is therefore crucial for understanding how diseases can result from failures in these systems. The proposed research addresses the question of how an oligomeric DNA-binding protein, which can in principle adopt a highly symmetric conformation, binds to a DNA recognition element that can be at most twofold symmetric. The tetrameric tumor suppresser p53, the trimeric heat shock transcription factor HSF, and the tetrameric lactose repressor are well-studied examples of such oligomeric proteins. In each case, the protein uses multiple DNA-binding domains to bind sequences composed of direct and inverted repeats of a recognition element. In the proposed research, the hexameric arginine repressor (ArgR) will serve as a model system for studying the architecture of an oligomeric protein-DNA assembly. ArgR plays a multifuctional role in the bacterial cell, serving as the master regulator of the arginine biosynthetic genes as well as playing an obligatory architectural role in mediating site-specific recombination events. In the presence of the corepressor L-arginine, ArgR uses four of its six DNA-binding domains to recognize DNA operators composed of a tandem repeat of palindromic """"""""Arg boxes"""""""". The ArgR DNA-binding domains are members of the winged helix-turn-helix (wHTH) family of DNA-binding motifs. This growing family includes domains found in histone H5, the ets family, HNF- 3/forkhead, and the heat shock transcription factor. In order to build a structural framework for understanding the basic principles of how oligomeric proteins recognize complex DNA sequences, and how multiple wHTH domains interact at the DNA surface, X-ray diffraction methods will be used to determine structures of (i) ArgR in the low affinity DNA-binding state, (ii) ArgR in the high affinity state with bound corepressor, (iii) an ArgR superrepressor, (iv) an ArgR-DNA complex representative of the architectural role of ArgR in site-specific recombination, and (v) an ArgR-DNA complex formed with a tandem Arg box operator.
| Wong, Ming-Ching; Dobi, Krista C; Baylies, Mary K (2014) Discrete levels of Twist activity are required to direct distinct cell functions during gastrulation and somatic myogenesis. PLoS One 9:e99553 |
| Gordi, Toufigh; Xie, Rujia; Jusko, William J (2005) Semi-mechanistic pharmacokinetic/pharmacodynamic modelling of the antimalarial effect of artemisinin. Br J Clin Pharmacol 60:594-604 |
