Kinetic Analysis as a Tool to Distinguish Pathway Complexity in Molecular Assembly: An Unexpected Outcome of Structures in Competition.
While the sensitive dependence of the functional characteristics of self-assembled nanofibers on the molecular structure of their building blocks is well-known, the crucial influence of the dynamics of the assembly process is often overlooked. For natural protein-based fibrils, various aggregation mechanisms have been demonstrated, from simple primary nucleation to secondary nucleation and off-pathway aggregation. Similar pathway complexity has recently been described in synthetic supramolecular polymers and has been shown to be intimately linked to their morphology. We outline a general method to investigate the consequences of the presence of multiple assembly pathways, and show how kinetic analysis can be used to distinguish different assembly mechanisms. We illustrate our combined experimental and theoretical approach by studying the aggregation of chiral bipyridine-extended 1,3,5-benzenetricarboxamides (BiPy-1) in n-butanol as a model system. Our workflow consists of nonlinear least-squares analysis of steady-state spectroscopic measurements, which cannot provide conclusive mechanistic information but yields the equilibrium constants of the self-assembly process as constraints for subsequent kinetic analysis. Furthermore, kinetic nucleation-elongation models based on one and two competing pathways are used to interpret time-dependent spectroscopic measurements acquired using stop-flow and temperature-jump methods. Thus, we reveal that the sharp transition observed in the aggregation process of BiPy-1 cannot be explained by a single cooperative pathway, but can be described by a competitive two-pathway mechanism. This work provides a general tool for analyzing supramolecular polymerizations and establishing energetic landscapes, leading to mechanistic insights that at first sight may seem unexpected and counterintuitive.
Institute for Complex Molecular Systems, ‡Laboratory of Macromolecular and Organic Chemistry, and §Computational Biology Group, Eindhoven University of Technology , P.O. Box 513, 5600 MB Eindhoven, The Netherlands.
Daan
D
van der Zwaag
Institute for Complex Molecular Systems, ‡Laboratory of Macromolecular and Organic Chemistry, and §Computational Biology Group, Eindhoven University of Technology , P.O. Box 513, 5600 MB Eindhoven, The Netherlands.
Pascal A
PA
Pieters
Institute for Complex Molecular Systems, ‡Laboratory of Macromolecular and Organic Chemistry, and §Computational Biology Group, Eindhoven University of Technology , P.O. Box 513, 5600 MB Eindhoven, The Netherlands.
Peter A
PA
Korevaar
Institute for Complex Molecular Systems, ‡Laboratory of Macromolecular and Organic Chemistry, and §Computational Biology Group, Eindhoven University of Technology , P.O. Box 513, 5600 MB Eindhoven, The Netherlands.
Albert J
AJ
Markvoort
Institute for Complex Molecular Systems, ‡Laboratory of Macromolecular and Organic Chemistry, and §Computational Biology Group, Eindhoven University of Technology , P.O. Box 513, 5600 MB Eindhoven, The Netherlands.
A J H
AJ
Spiering
Institute for Complex Molecular Systems, ‡Laboratory of Macromolecular and Organic Chemistry, and §Computational Biology Group, Eindhoven University of Technology , P.O. Box 513, 5600 MB Eindhoven, The Netherlands.
Tom F A
TF
de Greef
Institute for Complex Molecular Systems, ‡Laboratory of Macromolecular and Organic Chemistry, and §Computational Biology Group, Eindhoven University of Technology , P.O. Box 513, 5600 MB Eindhoven, The Netherlands.
E W
EW
Meijer
D020001:1-Butanol / Q000737:chemistry*; D002942:Circular Dichroism; D057927:Hydrophobic and Hydrophilic Interactions; D007700:Kinetics; D008958:Models, Molecular; D015394:Molecular Structure; D011725:Pyridines / Q000737:chemistry*; D013696:Temperature
D011725:Pyridines; D020001:1-Butanol; C023666:pyridine