Poster abstracts

Poster number 16 submitted by Ian Kleckner

Donut Dynamics: a Relaxation Dispersion Study of Allosteric Gene Regulation by the Ring-Protein TRAP

Ian R. Kleckner (Biophysics Program, The Ohio State University), Craig A. McElroy (The Ohio State University), Joseph R. Sachleben (The Ohio State University), Paul Gollnick (SUNY at Buffalo), Philipp Neudecker, Lewis E. Kay (University of Toronto), Mark P. Foster (Biochemistry Dept., The Ohio State University)

Abstract:
The tryptophan biosynthetic pathway in bacilli is regulated by the trp-RNA-binding Attenuation Protein (TRAP). TRAP forms a 90 kDa radially symmetric donut-shaped 11-mer which exhibits reduced dynamics upon binding 11 tryptophan (Trp) molecules. Activated Trp-TRAP binds a specific RNA sequence with high affinity and thus inhibits further transcription of the downstream genes.

This work will test the hypotheses that (1) apo TRAP's internal motions (dynamics) mask its RNA-binding surface and (2) Trp-binding activates TRAP for RNA-binding via changes in both structure and dynamics.

NMR spectroscopy is used to quantify site-specifc dynamics and structural changes of TRAP upon binding Trp. This study indicates Trp-binding abolishes the μs-ms dynamics measured in apo TRAP. Furthermore, probed regions of TRAP whose structure and dynamics are most sensitive to Trp-binding are limited to the Trp- and RNA-binding regions.

These significant findings provide quantitative insight into the rugged conformational landscape sampled by apo TRAP at the μs-ms timescale (in contrast, Trp-TRAP occupies a single well-defined state at this timescale). Moreover, the dynamics of apo TRAP are limited to where it is functionally relevant for masking the RNA-binding surface and permitting entry of Trp ligand.

More generally, this supports a minimalist model of allosteric regulation in which the allosteric regulator alters only a “hot spot” in the target protein to achieve the desired response.

Keywords: Protein dynamics, Allosteric regulation, NMR