2017 David F. Silbert Summer Fellowship Research

Divya Natarajan
2017 Summer Silbert Fellow
Summer Research Program Abstract
Divya Natarajan
David F. Silbert Summer Fellowship
Summer Research Program
Current Doctoral Program of Study: Medicine
Washington University in St. Louis
Department: Biomedical Engineering
Mentors: Rohit Pappu, PhD; Ammon Posey, PhD

Toward Rational Design of Huntington’s Disease Therapeutics Using Profilin as a Model of Huntingtin Aggregation Suppression Via Multivalent Interactions

Natarajan DK; Posey AE; Pappu RV

Introduction: Huntington’s disease (HD) is characterized by neuronal degeneration caused by fibrillar aggregates of N-terminal fragments of Huntingtin (Htt-NTF) with expanded glutamine (Q) repeats. Profilin is able to suppress aggregation of mutant huntingtin by stabilizing the soluble form of Htt-NTF and interacts with the polyproline (polyP) tracts in a region adjacent to the expanded polyQ region, which we term C38. Interestingly, our previous experiments revealed that the apparent affinity of profilin for QnC38 is higher than for C38 alone, and our computational results suggest that auxiliary interactions between profilin and polyQ may explain this increased affinity. We hypothesize that an auxiliary interaction between profilin and the polyQ region of Htt-NTFs supports the primary interaction with polyP tracts, and that this multivalency is important for the aggregation-reducing properties of profilin.

Methods: 15N-labeled profilin was recombinantly expressed and purified using previously established protocols. C38 and Q40C38 peptides were obtained in crude form from a commercial source and were purified in-house using RP-HPLC chromatography. 1H-15N HSQC NMR spectra were collected for 15N-labeled profilin in the presence of two different Htt-NTF peptide ligands, C38 and Q40C38, in order to identify the specific residues involved in the interaction between these molecules.

Results: The chemical shifts measured for profilin in the presence of C38 were consistent with those previously observed for profilin/P10, thereby confirming the interaction between the proline-rich C38 region of Htt-NTFs and the polyP binding site on profilin. The profilin/C38 spectrum also serves as a point of comparison for 1H-15N HSQCs measured in the presence of Q40C38 peptide, as any additional changes in the chemical shifts that are detected can be attributed to the hypothesized auxiliary interaction between profilin and the polyQ region of Htt-NTF. These results are pending.

Conclusions: Understanding the exact sites of interaction between profilin and Htt-NTF will facilitate rational drug development in order to synthesize a therapeutic that mimics the aggregation-reducing effects of profilin.