Nucleation‐Supersaturation Dual‐Drive Crystallization Strategy Enables Efficient Protein Crystallization

• Published in: Small (Weinheim an der Bergstrasse, Germany) Vol. 20; no. 20; pp. e2307924 - n/a
• Main Authors: Yan, Yizhen, Wang, Junyou, Lu, Xuechun, Yuan, Weikang, Zhang, Xiangyang
• Format: Journal Article
• Language: English
• Published: Germany Wiley Subscription Services, Inc 01.05.2024
• Subjects: Crystallization; Crystals; faster crystallization; high‐quality crystals; hydrogel template; Hydrogels; Lysozyme; Nucleation; nucleation‐supersaturation regulation; Protein crystallization; Proteins; Supersaturation; hydrogel template; nucleation-supersaturation regulation; Protein crystallization; faster crystallization; high-quality crystals
• ISSN: 1613-6810; 1613-6829
• DOI: 10.1002/smll.202307924

A rational crystallization strategy is essential to obtain high‐quality protein crystals, yet the established methods suffer from different limitations arising from the single regulation on either nucleation or supersaturation. Herein, a nucleation‐supersaturation dual‐driven crystallization (DDC) strategy that realizes synergistic regulation of heterogeneous nucleation sites and solution supersaturation based on dual surface and confinement effects for efficient protein crystallization is reported. This strategy relies on a p(PEGDA‐co‐DMAA) hydrogel template with pre‐filled NaCl under designed concentrations. Once dropping hen egg white lysozyme (HEWL) protein solution on the hydrogel, the wrinkled surface provides numerous nucleation sites, while the internal structure regulates the solution supersaturation in the crystallization region through diffusion. Finally, DDC strategy can create high‐quality HEWL crystals with large sizes (100–300 µm), well‐defined morphologies (hexagon and tetragon), and a significantly accelerated nucleation time (9–12 times faster than that achieved using the conventional hanging drop method). It also performs well at wider protein concentrations (10–50 mg mL−1) and categories (e.g., achieving fast crystallization and large‐size crystals of trypsin), therefore demonstrating clear advantages and great potential for efficiently fabricating protein crystals desirable for diverse applications. An innovative strategy for efficient protein crystallization is developed by realizing a synergistic regulation of nucleation sites and protein solution supersaturation, which relies on a p(PEGDA‐co‐DMAA) hydrogel template with pre‐filled NaCl. High‐quality hen egg white lysozyme (HEWL) and trypsin crystals with large size, well‐defined morphology, and accelerated nucleation time are obtained.