Concentration-Driven Assembly and Sol-Gel Transition of π-Conjugated Oligopeptides

Abstract

Advances in supramolecular assembly have enabled the design and synthesis of functional materials with well-defined structures across multiple length scales. Biopolymer-synthetic hybrid materials can assemble into supramolecular structures with a broad range of structural and functional diversity through precisely controlled noncovalent interactions between subunits. Despite recent progress, there is a need to understand the mechanisms underlying the assembly of biohybrid/synthetic molecular building blocks, which ultimately control the emergent properties of hierarchical assemblies. In this work, we study the concentration-driven self-assembly and gelation of π-conjugated synthetic oligopeptides containing different π-conjugated cores (quaterthiophene and perylene diimide) using a combination of particle tracking microrheology, confocal fluorescence microscopy, optical spectroscopy, and electron microscopy. Our results show that π-conjugated oligopeptides self-assemble into β-sheet-rich fiber-like structures at neutral pH, even in the absence of electrostatic screening of charged residues. A critical fiber formation concentration c_fiber and a critical gel concentration c_gel are determined for fiber-forming π-conjugated oligopeptides, and the linear viscoelastic moduli (storage modulus G' and loss modulus G″) are determined across a wide range of peptide concentrations. These results suggest that the underlying chemical structure of the synthetic π-conjugated cores greatly influences the self-assembly process, such that oligopeptides appended to π-conjugated cores with greater torsional flexibility tend to form more robust fibers upon increasing peptide concentration compared to oligopeptides with sterically constrained cores. Overall, our work focuses on the molecular assembly of π-conjugated oligopeptides driven by concentration, which is controlled by a combination of enthalpic and entropic interactions between oligopeptide subunits.

Figure 1

Investigating the self-assembly of π-conjugated oligopeptides at neutral pH. (a) Chemical structures of DFAG-OT4 and DFAG-PDI. (b) Schematic of the experimental setup for multiple particle tracking microrheology (PTM). (c) Characteristic image of tracer particles (diameter d = 0.84 μm) diffusing in a peptide matrix from fluorescence microscopy.

Figure 2

Microrheology of the sol–gel transition for DFAG-OT4. (a) Ensemble-averaged MSDs ⟨Δr²(τ)⟩ versus lag time τ for probe particles in DFAG-OT4 solutions at different peptide concentrations. The dashed line shows the slope for the critical diffusive exponent α_crit = 0.78. (b) Time-cure superposition (TCS) of the MSD for DFAG-OT4 as a function of oligopeptide concentration. The master curve converges with a critical exponent α_crit = 0.78 ± 0.04 at the critical gel concentration c_gel = 1 mg/mL.

Figure 3

Assembly of DFAG-OT4 using a combination of microrheology, optical spectroscopy, and cryo-electron microscopy. (a) Diffusive exponent α from MSDs and peak fluorescence emission wavelength as functions of peptide concentration for DFAG-OT4. Error bars for the diffusive exponent denote the standard deviation from multiple measurements. (b) CD spectra of DFAG-OT4 as a function of peptide concentration. (c) Confocal fluorescence microscopy images of DFAG-OT4 droplets at different peptide concentrations. Arrows indicate the edge of the droplet. (d) Cryo-SEM image of freeze-dried DFAG-OT4 solution at 2 mg/mL.

Figure 4

Viscoelastic moduli G′(ω) and G″(ω) as a function of frequency ω determined from microrheology experiments for DFAG-OT4 at neutral pH (pH = 7) at (a) 0.5 mg/mL and (b) 2 mg/mL. (c) Representative probe particle trajectories and (d) van Hove correlation functions of probe particle trajectories at peptide concentrations of 0.1 mg/mL, 1 mg/mL, and 2 mg/mL. Solid lines are Gaussian fits. Scale bar = 2 μm.

Figure 5

Assembly of DFAG-PDI using a combination of microrheology, optical spectroscopy, and cryo-electron microscopy. (a) Ensemble-averaged MSDs ⟨Δr²(τ)⟩ as a function of lag time τ for probe particles in DFAG-PDI solutions with different concentrations. (b) Diffusive exponent α from microrheology and peak fluorescence emission wavelength as functions of peptide concentration for DFAG-PDI. Error bars for the diffusive exponent denote the standard deviation from multiple measurements. (c) Confocal fluorescence microscopy images of DFAG-PDI droplets at different concentrations. Arrow indicate the edge of the droplet. (d) Cryo-SEM image of DFAG-PDI solution at 1 mg/mL.