Cosolute effects reveal the nature of weak forces governing GLP-1 oligomer stability

TL;DR

This study explores how GLP-1 hormone forms stable oligomers and how cosolutes affect their aggregation into fibrils, which is important for drug development.

Contribution

The study reveals the role of weak forces and cosolutes in GLP-1 oligomer stability and aggregation mechanisms under pH 5 conditions.

Findings

GLP-1am forms soluble oligomers that assemble into nanosheets over hours in quiescent conditions.

Aggregation of GLP-1am is nucleation-dependent and accelerated by ionic cosolutes due to electrostatic screening.

Proline delays aggregation of GLP-1am in a pH-dependent manner.

Abstract

Glucagon-like peptide-1 (GLP-1) is an incretin hormone widely used to manage diabetes and obesity through its ability to regulate glucose homeostasis. Clinically relevant GLP-1 sequences form oligomeric states. Uncontrolled oligomer formation can drive fibril formation, posing challenges, such as difficulty in controlling drug dosage, loss of activity, or toxicity, as the aggregates can be immunogenic and/or can form amyloids. Here, we used combined measurements of colloidal and conformational stability to characterize the intermolecular interactions underpinning the physical status of the GLP-1(7–37) amide (GLP-1am), at pharmaceutically relevant high concentrations. We focus on less explored conditions, around pH 5, mimicking the environment within native cellular secretory granules, where the hormone is also densely packed. Cosolutes allowed us to interfere with weak interactions affecting peptide self-association into soluble oligomers and the conversion into aggregates and fibrils. We show that GLP-1am exists as soluble oligomers that assemble into nanosheets over the timescale of hours, in quiescent conditions. Aggregation proceeded via a nucleation-dependent mechanism, with its rate correlating to the magnitude of attractive intermolecular interactions. It was accelerated by ionic cosolutes, indicating a key role for screening of electrostatic interactions in modulating peptide–peptide attraction and assembly. The rate of aggregation was also pH dependent, with rates being slower at pH 5 than pH 8. Notably, the addition of proline, as a cosolute, delayed the onset of GLP-1am aggregation in a pH-dependent manner. Thus, in quiescent conditions, GLP-1am forms discrete soluble oligomers capable of organizing into ordered nanostructures rather than amyloid fibrils.

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