Giannetti, G. | bioRxiv Science

bioRxiv2026

Introducing non-enzymatic crosslinks into atomistic simulations of collagen fibrils

MotivationCollagen fibrils are the primary load-bearing units of connective tissues. However, generating atomistic, simulation-ready models remains challenging due to collagens hierarchical organization and the diversity of its crosslinking network across tissues, ages, and metabolic states. Notably, non-enzymatic advanced glycation end-product (AGE) crosslinks--central to aging and diabetic complications--are largely absent from current atomistic fibril modelling workflows. ResultsHere, we present an extension of the ColBuilder framework to generate atomistic collagen fibril models that incorporate three representative AGE-derived crosslinks (glucosepane, pentosidine, and MOLD) alongside enzymatic crosslinks. Amber99-compatible parameters are provided and assessed against QM-optimized reference geometries using all-atom molecular dynamics (MD) simulations. As proof-of-concept, we examine the mechanical response of single D-period collagen microfibrils featuring enzymatic-only, AGE-only, and mixed crosslink patterns in Molecular Dynamics simulations under force, and observe that AGE crosslinks differently impact the fibril structure compared to enzymatic crosslinks. The extension to ColBuilder can aid future structure-based research on collagen aging. Availability and implementationColBuilder is available as an open-source Python command-line package at https://github.com/graeter-group/colbuilder.

bioinformatics↗

bioRxiv2023

Elucidating the role of water in collagen self assembly by isotopically modulating collagen hydration

Water is known to play an important role in collagen self assembly, but it is still largely unclear how water-collagen interactions influence the assembly process and determine the fibril network properties. Here, we use the H2O/D2O isotope effect on the hydrogen-bond strength in water to investigate the role of hydration in collagen self assembly. We dissolve collagen in H2O and D2O, and compare the growth kinetics and the structure of the collagen assemblies formed in these water isotopomers. Surprisingly, collagen assembly occurs ten times faster in D2O than in H2O, and collagen in D2O self assembles into much thinner fibrils, that form a more inhomogeneous and softer network, with a fourfold reduction in elastic modulus compared to H2O. Combining spectroscopic measurements with atomistic simulations, we show that collagen in D2O is less hydrated than in H2O. This partial dehydration lowers the enthalpic penalty for water removal and reorganization at the collagen-water interface, increasing the self assembly rate and the number of nucleation centers, leading to thinner fibrils and a softer network. Coarse-grained simulations show that the acceleration in the initial nucleation rate can be reproduced by the enhancement of electrostatic interactions, which appear to be crucial in determining the acceleration of the initial nucleation rate. These results show that water acts as a mediator between collagen monomers, by moderating their interactions so as to optimize the assembly process and, thus, the final network properties. We believe that isotopically modulating the hydration of proteins can be a valuable method to investigate the role of water in protein structural dynamics and protein self assembly.

biophysics↗