Preparation of Mechanically Stable Self-Assembled Peptides Hydrogels

Summary

This protocol presents three fast and simple preparation methods that use environmental conditions to trigger the self-assembly of peptides into hydrogels. Additionally, the characterization of peptide hydrogels is described, demonstrating that mechanically stable peptide hydrogels can be formed under these straightforward conditions.

Abstract

Peptide hydrogels are highly hydrophilic, three-dimensional network gels formed by the self-assembly of nanofibers or polymers, creating water-locking networks. Their morphology closely resembles that of the extracellular matrix, allowing them to exhibit both the biological functions of peptides and responsive gelation properties. These unique characteristics have led to their extensive application in tissue engineering, three-dimensional cell culture, cancer therapy, regenerative medicine, and other biomedical fields. This article describes three methods for preparing ECF-5 peptide hydrogels using self-assembling peptides with environmentally responsive gelation processes: (1) pH-responsive gelation: varying pH levels induce the protonation or deprotonation of amino acid residues, altering electrostatic interactions between peptide molecules and promoting their self-assembly into hydrogels; (2) Metal ion addition: polyvalent metal ions chelate with negatively charged amino acid residues, acting as bridges between peptides to form a network hydrogel; (3) Solvent exchange: hydrophobic peptides are initially dissolved in non-polar organic solvents and subsequently induce self-assembly into hydrogels upon transitioning to a polar aqueous environment. These methods utilize conventional experimental procedures to facilitate peptide self-assembly into hydrogels. By designing peptide sequences to align with specific gelation-inducing conditions, it is possible to achieve finely tuned micro/nanostructures and biological functions, highlighting the significant potential of peptide hydrogels in the biomedical domain.

Introduction

Through the design of peptide sequences, non-covalent interactions between peptides induce self-assembly, leading to the formation of ordered micro- and nanometer structures, including nanotubes, nanoribbons, nanofibers, and spherical structures¹. When self-assembled into micro- and nanometer fibers/ribbons, these structures macroscopically exhibit hydrogel properties. Peptide self-assembling hydrogels differ from polymer hydrogels in that they self-assemble through non-covalent interactions, their gel form is reversible, and they readily respond to specific conditions to transition between solution and gel phases². For ....

Protocol

The details of the plasmids, reagents, and equipment used in this study are listed in the Table of Materials.

1. pH response method

1. Add 5 mg of ECF-5 peptides to 400 µL of deionized water. Sonicate at 40 kHz for 30 min and mix thoroughly.
2. Add 40 µL of sodium hydroxide (1 M, filtered through a 0.22 µm filter) to the peptide solution. Vortex and mix thoroughly. Continue sonication for 15 min until the solution is completely c.......

Discussion

In the past few decades, following the discovery of self-assembling peptide sequences derived from amyloid proteins, numerous self-assembling peptides have been designed based on their properties, demonstrating significant potential for applications in biomedicine and materials science¹⁹. Peptide hydrogels have exhibited unique bio-functionalization capabilities in tissue culture, drug delivery, and tumor treatment²⁰.

This article describes simpl.......

Materials

Name	Company	Catalog Number	Comments
3-Aminopropyl)triethoxysilane	Aladdin	A107147	/
Atomic Force Microscopy	Bruker	Multimode Nanoscope VIII	/
CaCl2	Aladdin	C290953	/
Diphenylalanine (FF)	Chinesepeptide	customizable	Purity > 95%
DMSO	Sigma-aldrich	34869	/
ECF-5 Peptides	Chinesepeptide	sequence: ECAFF	Purity > 95%
Hydrochloric Acid	Aladdin	H399657	/
Mica	Sigma-aldrich	AFM-71856-02	/
Phosphate Buffered Saline	Aladdin	P492453	/
Rheometer	Anton Paar GmbH	MCR302	/
Silicon Cantilevers	MikroMasch	XSC11	/
Sodium Chloride	Aladdin	C111549	/
Sodium Hydroxide	Aladdin	S140903	/
TRIS Hydrochloride	Aladdin	T431531	/