Interaction of Huntingtin Exon-1 Peptides with Lipid-Based Micellar Nanoparticles Probed by Solution NMR and Q-Band Pulsed EPR

Abstract

Lipid-based micellar nanoparticles promote aggregation of huntingtin exon-1 peptides. Here we characterize the interaction of two such peptides, httNTQ7 and httNTQ10 comprising the N-terminal amphiphilic domain of huntingtin followed by 7 and 10 glutamine repeats, respectively, with 8 nm lipid micelles using NMR chemical exchange saturation transfer (CEST), circular dichroism and pulsed Q-band EPR. Exchange between free and micelle-bound httNTQn peptides occurs on the millisecond time scale with a KD ~ 0.5–1 mM. Upon binding micelles, residues 1–15 adopt a helical conformation. Oxidation of Met7 to a sulfoxide reduces the binding affinity for micelles ~3–4-fold and increases the length of the helix by a further two residues. A structure of the bound monomer unit is calculated from the backbone chemical shifts of the micelle-bound state obtained from CEST. Pulsed Q-band EPR shows that a monomer–dimer equilibrium exists on the surface of the micelles and that the two helices of the dimer adopt a parallel orientation, thereby bringing two disordered polyQ tails into close proximity which may promote aggregation upon dissociation from the micelle surface.

The folding equilibrium of huntingtin exon 1 monomer depends on its polyglutamine tract

“…The difference in monomer structures may also impact its aggregation propensities. The aggregation of Httex1 proceeds via a stepwise mechanism that is initiated by an oligomer predominantly stabilized by ␣-helical structure in the N-terminal region in solution and on the membrane (37,43,44). Moreover, Wetzel and co-workers found concentration-dependent helicity in the N17 (45,46), signifying that this region becomes more helical with oligomerization.…”

Decorrelating Kinetic and Relaxation Parameters in Exchange Saturation Transfer NMR: A Case Study of N-Terminal Huntingtin Peptides Binding to Unilamellar Lipid Vesicles

“…12,13 Recently, we showed, using chemical exchange saturation transfer (CEST) NMR and Q-band-pulsed electron paramagnetic resonance (EPR), that both peptides form a well-defined α -helical structure when bound to lipid micelles and, in addition, can dimerize in the bound state. 14 As small unilamelar lipid vesicles (SUV) have a molecular weight of ∼4.3 MDa and are ∼4-fold larger in diameter than micelles (∼31 versus ∼8 nm), the same experimental approach cannot be used to study the exchange of htt NT and htt NT Q 7 with their SUV-bound form, and hence, experiments such as DEST and Δ R 2 are required. However, in the exchange regime where the transverse spin relaxation rate R2B of the bound state is smaller than the strength of the RF saturation field, DEST and Δ R 2 data are only sensitive to the product of R2B and the population p B of the bound state, and hence, R2B and p B cannot be determined independently.…”

TiO2 Nanoparticles Catalyze Oxidation of Huntingtin Exon 1-Derived Peptides Impeding Aggregation: A Quantitative NMR Study of Binding and Kinetics

“…The 15 N-DEST/Δ R 2 data for Leu 3 and the C-terminal three residues of htt NT required a separate treatment and were fitted to the three-state model shown in gray in Figure 3B. Although formally the same overall three-state exchange model is used for these residues PFred ↔ PRred ↔ PTred, the exchange process subsumes initial binding to the NP surface followed by reversible detachment from the surface (state PTred, where “T” denotes “tethered”).…”