Classical and Nonclassical Nucleation Mechanisms of Insulin Crystals

• Type: Article in International Scientific Journal
• Date: 2024-04-08

Authors

Campos, JBLM (Author) FEUP

Journal

Title: ACS Omega

Vol. 9 No. 6

Publisher: American Chemical Society

Indexing

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Other information

Authenticus ID: P-010-EYV

DOI: 10.1021/acsomega.3c10052

Abstract (EN): Although the Classical Nucleation Theory (CNT) is the most consensual theory to explain protein nucleation mechanisms, experimental observations during the shear-induced assays suggest that the CNT does not always describe the insulin nucleation process. This is the case at intermediate precipitant (ZnCl2) solution concentrations (2.3 mM) and high-temperature values (20 and 40 degrees C) as well as at low precipitant solution concentrations (1.6 mM) and low-temperature values (5 degrees C). In this work, crystallization events following the CNT registered at high precipitant solution concentrations (3.1 and 4.7 mM) are typically described by a Newtonian response. On the other hand, crystallization events following a nonclassical nucleation pathway seem to involve the formation of a metastable intermediate state before crystal formation and are described by a transition from Newtonian to shear-thinning responses. A dominant shear-thinning behavior (shear viscosity values ranging more than 6 orders of magnitude) is found during aggregation/agglomeration events. The rheological analysis is complemented with different characterization techniques (Dynamic Light Scattering, Energy-Dispersive Spectroscopy, Circular Dichroism, and Differential Scanning Calorimetry) to understand the insulin behavior in solution, especially during the occurrence of aggregation/agglomeration events. To the best of our knowledge, the current work is the first study describing nonclassical nucleation mechanisms during shear-induced crystallization experiments, which reveals the potential of the interdisciplinary approach herein described and opens a window for a clear understanding of protein nucleation mechanisms.

Language: English

Type (Professor's evaluation): Scientific

No. of pages: 13