The Internal Structural Dynamics of Elastin-Like Polypeptide Assemblies by 13C-Direct Detected NMR Spectroscopy

Abstract

Elastin-like polypeptides (ELPs) are biocompatible polymers exhibiting lower critical solution temperature (LCST) behavior, making them valuable in various applications, including drug delivery and tissue engineering. This study addresses the atomistic-level understanding of ELP self-assembly, focusing on their internal structural dynamics. Conventional proton-detected nuclear magnetic resonance (NMR) spectroscopy faces limitations in studying ELP aggregates due to accelerated proton exchange processes, which cause significant resonance broadening. Herein, we show how to overcome this hurdle by using carbon-13-detected NMR. This method mitigates issues related to amide proton exchange, allowing for a residue-resolved view of the internal configuration of ELP aggregates. With this method, we record residue-resolved ¹⁵N relaxation rates, revealing three features. (i) Proline residues within the PGXGV pentapeptide repeats (X being any amino acid except proline) of ELP become motional restricted upon aggregation, indicating their role as interchain contacts. (ii) Pentapeptides with alanine guest residue X display particularly significantly reduced motional freedom upon aggregation. (iii) Even within large ELP aggregates, fast internal dynamics characterize the peptide chains in a way that is reminiscent of condensed liquid phases. The presented study is the first proof of concept that ¹³C-direct detection is a viable tool to delineate the internal structural dynamics of condensed ELP phases by NMR. It might, thus, help to foster new investigations of their aggregation mechanisms.