Hierarchical nanomechanics of collagen microfibrils

Collagen constitutes one third of the human proteome, providing mechanical stability, elasticity and strength to connective tissues. Collagen is also the dominating material in the extracellular matrix (ECM) and is thus crucial for cell differentiation, growth and pathology. However, fundamental questions remain with respect to the origin of the unique mechanical properties of collagenous tissues, and in particular its stiffness, extensibility and nonlinear mechanical response. By using x-ray diffraction data of a collagen fibril reported by Orgel et al. (Proceedings of the National Academy of Sciences USA, 2006) in combination with protein structure identification methods, here we present an experimentally validated model of the nanomechanics of a collagen microfibril that incorporates the full biochemical details of the amino acid sequence of the constituting molecules. We report the analysis of its mechanical properties under different levels of stress and solvent conditions, using a full-atomistic force field including explicit water solvent. Mechanical testing of hydrated collagen microfibrils yields a Young’s modulus of ≈300 MPa at small and ≈1.2 GPa at larger deformation in excess of 10% strain, in excellent agreement with experimental data. Dehydrated, dry collagen microfibrils show a significantly increased Young’s modulus of ≈1.8 to 2.25 GPa (or ≈6.75 times the modulus in the wet state) owing to a much tighter molecular packing, in good agreement with experimental measurements (where an increase of the modulus by ≈9 times was found). Our model demonstrates that the unique mechanical properties of collagen microfibrils can be explained based on their hierarchical structure, where deformation is mediated through mechanisms that operate at different hierarchical levels. Key mechanisms involve straightening of initially disordered and helically twisted molecules at small strains, followed by axial stretching of molecules, and eventual molecular uncoiling at extreme deformation. These mechanisms explain the striking difference of the modulus of collagen fibrils compared with single molecules, which is found in the range of 4.8±2 GPa or ≈10-20 times greater. These findings corroborate the notion that collagen tissue properties are highly scale dependent and nonlinear elastic, an issue that must be considered in the development of models that describe the interaction of cells with collagen in the extracellular matrix. A key impact the atomistic model of collagen microfibril mechanics reported here is that it enables the bottom-up elucidation of structure-property relationships in the broader class of collagen materials such as tendon or bone, including studies in the context of genetic disease where the incorporation of biochemical, genetic details in material models of connective tissue is essential

Nanostructure and stability of calcitonin amyloids

Calcitonin is a 32-amino acid thyroid hormone that can form amyloid fibrils. The structural basis of the fibril formation and stabilization is still debated and poorly understood. The reason is that NMR data strongly suggest antiparallel β-sheet calcitonin assembly, whereas modeling studies on the short DFNKF peptide (corresponding to the sequence from Asp15 to Phe19 of human calcitonin and reported as the minimal amyloidogenic module) show that it assembles with parallel β-sheets. In this work, we first predict the structure of human calcitonin through two complementary molecular dynamics (MD) methods, finding that human calcitonin forms an α-helix. We use extensive MD simulations to compare previously proposed calcitonin fibril structures. We find that two conformations, the parallel arrangement and one of the possible antiparallel structures (with Asp15 and Phe19 aligned), are highly stable and ordered. Nonetheless, fibrils with parallel molecules show bulky loops formed by residues 1 to 7 located on the same side, which could limit or prevent the formation of larger amyloids. We investigate fibrils formed by the DFNKF peptide by simulating different arrangements of this amyloidogenic core sequence. We show that DFNKF fibrils are highly stable when assembled in parallel β-sheets, whereas they quickly unfold in antiparallel conformation. Our results indicate that the DFNKF peptide represents only partially the full-length calcitonin behavior. Contrary to the full-length polypeptide, in fact, the DFNKF sequence is not stable in antiparallel conformation, suggesting that the residue flanking the amyloidogenic peptide contributes to the stabilization of the experimentally observed antiparallel β-sheet packing

Unbiased in silico design of pH-sensitive tetrapeptides

We used coarse-grain molecular dynamics simulations to screen all possible histidine-bearing tetrapeptide sequences, finding novel peptide sequences with pH-tunable assembly properties. These tetrapeptides could be used for various biological applications, such as triggered delivery of bioactive molecules

Molecular dynamics simulations provide insights into the substrate specificity of FAOX family members

Enzymatic assays based on Fructosyl Amino Acid Oxidases (FAOX) represent a potential, rapid and economical strategy to measure glycated hemoglobin (HbA1c), which is in turn a reliable method to monitor the insurgence and the development of diabetes mellitus. However, the engineering of naturally occurring FAOX to specifically recognize fructosyl-valine (the glycated N-terminal residue of HbA1c) has been hindered by the paucity of information on the tridimensional structures and catalytic residues of the different FAOX that exist in nature, and in general on the molecular mechanisms that regulate specificity in this class of enzymes. In this study, we use molecular dynamics simulations and advanced modeling techniques to investigate five different relevant wild-type FAOX (Amadoriase I, Amadoriase II, PnFPOX, FPOX-E and N1-1-FAOD) in order to elucidate the molecular mechanisms that drive their specificity towards polar and nonpolar substrates. Specifically, we compare these five different FAOX in terms of overall folding, ligand entry tunnels, ligand binding residues and ligand binding energies. Our work will contribute to future enzyme structure modifications aimed at the rational design of novel biosensors for the monitoring of blood glucose levels

Molecular dynamics simulations of the intrinsically disordered protein amelogenin

Amelogenin refers to a class of intrinsically disordered proteins that are the major constituents of enamel matrix derivative (EMD), an extract of porcine fetal teeth used in regenerative periodontal therapy. Modifications in molecular conformation induced by external stresses, such as changes in temperature or pH, are known to reduce the effectiveness of EMD. However, detailed descriptions of the conformational behavior of native amelogenin are lacking in the open literature. In the present work, a molecular model for the secondary and tertiary structure of the full-length major porcine amelogenin P173 was constructed from its primary sequence by replica exchange molecular dynamics (REMD) simulations. The REMD results for isolated amelogenin molecules at different temperatures were shown to be consistent with the available spectroscopic data. They therefore represent an important first step toward the simulation of the intra- and intermolecular interactions that mediate self-organization in amelogenin and its behavior in the presence of other EMD components under conditions representative of its therapeutic application

In Silico Analysis of Nanoplastics' and β-amyloid Fibrils' Interactions

: Plastic pollution has become a global environmental threat, which leads to an increasing concern over the consequences of plastic exposition on global health. Plastic nanoparticles have been shown to influence the folding of proteins and influence the formation of aberrant amyloid proteins, therefore potentially triggering the development of systemic and local amyloidosis. This work aims to study the interaction between nanoplastics and β-amyloid fibrils to better understand the potential role of nanoplastics in the outbreak of neurodegenerative disorders. Using microsecond-long coarse-grained molecular dynamics simulations, we investigated the interactions between neutral and charged nanoparticles made of the most common plastic materials (i.e., polyethylene, polypropylene, and polystyrene) and β-amyloid fibrils. We observe that the occurrence of contacts, region of amyloid fibril involved, and specific amino acids mediating the interaction depend on the type and charge of the nanoparticles

Advanced glycation end-products: Mechanics of aged collagen from molecule to tissue

Concurrent with a progressive loss of regenerative capacity, connective tissue aging is characterized by a progressive accumulation of Advanced Glycation End-products (AGEs). Besides being part of the typical aging process, type II diabetics are particularly affected by AGE accumulation due to abnormally high levels of systemic glucose that increases the glycation rate of long-lived proteins such as collagen. Although AGEs are associated with a wide range of clinical disorders, the mechanisms by which AGEs contribute to connective tissue disease in aging and diabetes are still poorly understood. The present study harnesses advanced multiscale imaging techniques to characterize a widely employed . in vitro model of ribose induced collagen aging and further benchmarks these data against experiments on native human tissues from donors of different age. These efforts yield unprecedented insight into the mechanical changes in collagen tissues across hierarchical scales from molecular, to fiber, to tissue-levels. We observed a linear increase in molecular spacing (from 1.45. nm to 1.5. nm) and a decrease in the D-period length (from 67.5. nm to 67.1. nm) in aged tissues, both using the ribose model of . in vitro glycation and in native human probes. Multiscale mechanical analysis of . in vitro glycated tendons strongly suggests that AGEs reduce tissue viscoelasticity by severely limiting fiber-fiber and fibril-fibril sliding. This study lays an important foundation for interpreting the functional and biological effects of AGEs in collagen connective tissues, by exploiting experimental models of AGEs crosslinking and benchmarking them for the first time against endogenous AGEs in native tissue