Reconstituted collagen fibrils. Fibrillar and molecular stability of the collagen upon maturation in vitro.
Ontology highlight
ABSTRACT:
SUBMITTER: Danielsen CC
PROVIDER: S-EPMC1144228 | biostudies-other | 1984 Sep
REPOSITORIES: biostudies-other
Similar Datasets
Project description:1. Interaction of bilirubin with collagen fibrils was explored in a two-phase system where collagen was present as an opaque rigid gel composed of striated fibrils, and bilirubin as an aqueous solution. 2. The Ka value of the binding of bilirubin to collagen fibrils is 5.4 X 10(3)M-1. The interaction of bilirubin with collagen fibrils depends on temperature. Below 5 degrees C, the binding is greatly diminished and denaturation of collagen fibril aggregates at 52--53 degrees C into a dissolution state abolishes binding of bilirubin. 3. Salicylate and sulphanilamide do not affect the binding of bilirubin to reconstituted collagen fibrils. 4. Serum albumin (40--80mM), known to reverse the binding of bilirubin to lipids, dissociates only 50% of the bilirubin bound to collagen fibrils. This suggests that sites located on collagen participate in some tight binding of bilirubin and the corresponding binding sites on albumin do not compete with them. 5. Urea (4M) abolishes more than 70% of the binding of bilirubin to collagen. Urea and thermal denaturation studies indicate the importance of conformation and organization of collagen fibrillar aggregates for the binding of bilirubin.
| S-EPMC1165431 | biostudies-other
Project description:Bone is a natural composite of collagen protein and the mineral hydroxyapatite. The structure of bone is known to be important to its load-bearing characteristics, but relatively little is known about this structure or the mechanism that govern deformation at the molecular scale. Here we perform full-atomistic calculations of the three-dimensional molecular structure of a mineralized collagen protein matrix to try to better understand its mechanical characteristics under tensile loading at various mineral densities. We find that as the mineral density increases, the tensile modulus of the network increases monotonically and well beyond that of pure collagen fibrils. Our results suggest that the mineral crystals within this network bears up to four times the stress of the collagen fibrils, whereas the collagen is predominantly responsible for the material's deformation response. These findings reveal the mechanism by which bone is able to achieve superior energy dissipation and fracture resistance characteristics beyond its individual constituents.
| S-EPMC3644085 | biostudies-literature
Project description:Collagen, a triple-helical, self-organizing protein, is the predominant structural protein in mammals. It is found in bone, ligament, tendon, cartilage, intervertebral disc, skin, blood vessel, and cornea. We have recently postulated that fibrillar collagens (and their complementary enzymes) comprise the basis of a smart structural system which appears to support the retention of molecules in fibrils which are under tensile mechanical strain. The theory suggests that the mechanisms which drive the preferential accumulation of collagen in loaded tissue operate at the molecular level and are not solely cell-driven. The concept reduces control of matrix morphology to an interaction between molecules and the most relevant, physical, and persistent signal: mechanical strain.The investigation was carried out in an environmentally-controlled microbioreactor in which reconstituted type I collagen micronetworks were gently strained between micropipettes. The strained micronetworks were exposed to active matrix metalloproteinase 8 (MMP-8) and relative degradation rates for loaded and unloaded fibrils were tracked simultaneously using label-free differential interference contrast (DIC) imaging. It was found that applied tensile mechanical strain significantly increased degradation time of loaded fibrils compared to unloaded, paired controls. In many cases, strained fibrils were detectable long after unstrained fibrils were degraded.In this investigation we demonstrate for the first time that applied mechanical strain preferentially preserves collagen fibrils in the presence of a physiologically-important mammalian enzyme: MMP-8. These results have the potential to contribute to our understanding of many collagen matrix phenomena including development, adaptation, remodeling and disease. Additionally, tissue engineering could benefit from the ability to sculpt desired structures from physiologically compatible and mutable collagen.
| S-EPMC2925882 | biostudies-literature
Project description:BackgroundAccording to the prevailing view, soluble oligomers or small fibrillar fragments are considered to be the most toxic species in prion diseases. To test this hypothesis, two conformationally different amyloid states were produced from the same highly pure recombinant full-length prion protein (rPrP). The cytotoxic potential of intact fibrils and fibrillar fragments generated by sonication from these two states was tested using cultured cells.Methodology/principal findingsFor one amyloid state, fibril fragmentation was found to enhance its cytotoxic potential, whereas for another amyloid state formed within the same amino acid sequence, the fragmented fibrils were found to be substantially less toxic than the intact fibrils. Consistent with the previous studies, the toxic effects were more pronounced for cell cultures expressing normal isoform of the prion protein (PrP(C)) at high levels confirming that cytotoxicity was in part PrP(C)-dependent. Silencing of PrP(C) expression by small hairpin RNAs designed to silence expression of human PrP(C) (shRNA-PrP(C)) diminished the deleterious effects of the two amyloid states to a different extent, suggesting that the role of PrP(C)-mediated and PrP(C)-independent mechanisms depends on the structure of the aggregates.Conclusions/significanceThis work provides a direct illustration that the relationship between an amyloid's physical dimension and its toxic potential is not unidirectional but is controlled by the molecular structure of prion protein (PrP) molecules within aggregated states. Depending on the structure, a decrease in size of amyloid fibrils can either enhance or abolish their cytotoxic effect. Regardless of the molecular structure or size of PrP aggregates, silencing of PrP(C) expression can be exploited to reduce their deleterious effects.
| S-EPMC3098877 | biostudies-literature
Project description:At the molecular scale, bone is mainly constituted of type-I collagen, hydroxyapatite, and water. Different fractions of these constituents compose different composite materials that exhibit different mechanical properties at the nanoscale, where the bone is characterized as a fiber, i.e., a bundle of mineralized collagen fibrils surrounded by water and hydroxyapatite in the extra-fibrillar volume. The literature presents only models that resemble mineralized collagen fibrils, including hydroxyapatite in the intra-fibrillar volume only, and lacks a detailed prescription on how to devise such models. Here, we present all-atom bone molecular models at the nanoscale, which, differently from previous bone models, include hydroxyapatite both in the intra-fibrillar volume and in the extra-fibrillar volume, resembling fibers in bones. Our main goal is to provide a detailed prescription on how to devise such models with different fractions of the constituents, and for that reason, we have made step-by-step scripts and files for reproducing these models available. To validate the models, we assessed their elastic properties by performing molecular dynamics simulations that resemble tensile tests, and compared the computed values against the literature (both experimental and computational results). Our results corroborate previous findings, as Young's Modulus values increase with higher fractions of hydroxyapatite, revealing all-atom bone models that include hydroxyapatite in both the intra-fibrillar volume and in the extra-fibrillar volume as a path towards realistic bone modeling at the nanoscale.
| S-EPMC8955169 | biostudies-literature
Project description:Collagen is the most abundant protein in the extracellular matrix (ECM), where its structural organization conveys mechanical information to cells. Using optical-tweezers-based microrheology, we investigated mechanical properties both of collagen molecules at a range of concentrations in acidic solution where fibrils cannot form and of gels of collagen fibrils formed at neutral pH, as well as the development of microscale mechanical heterogeneity during the self-assembly process. The frequency scaling of the complex shear modulus even at frequencies of ?10 kHz was not able to resolve the flexibility of collagen molecules in acidic solution. In these solutions, molecular interactions cause significant transient elasticity, as we observed for 5 mg/ml solutions at frequencies above ?200 Hz. We found the viscoelasticity of solutions of collagen molecules to be spatially homogeneous, in sharp contrast to the heterogeneity of self-assembled fibrillar collagen systems, whose elasticity varied by more than an order of magnitude and in power-law behavior at different locations within the sample. By probing changes in the complex shear modulus over 100-minute timescales as collagen self-assembled into fibrils, we conclude that microscale heterogeneity appears during early phases of fibrillar growth and continues to develop further during this growth phase. Experiments in which growing fibrils dislodge microspheres from an optical trap suggest that fibril growth is a force-generating process. These data contribute to understanding how heterogeneities develop during self-assembly, which in turn can help synthesis of new materials for cellular engineering.
| S-EPMC3732230 | biostudies-literature
Project description:According to a prevailing theory, (2S,4R)-4-hydroxyproline (Hyp) residues stabilize the collagen triple helix via a stereoelectronic effect that preorganizes appropriate backbone torsion angles for triple-helix formation. This theory is consistent with the marked stability that results from replacing the hydroxyl group with the more electron-withdrawing fluoro group, as in (2S,4R)-4-fluoroproline (Flp). Nonetheless, the hyperstability of triple helices containing Flp has been attributed by others to the hydrophobic effect rather than a stereoelectronic effect. We tested this hypothesis by replacing Hyp with 4,4-difluoroproline (Dfp) in collagen-related peptides. Dfp retains the hydrophobicity of Flp, but lacks the ability of Flp to preorganize backbone torsion angles. Unlike Flp, Dfp does not endow triple helices with elevated stability, indicating that the hyperstability conferred by Flp is not due to the hydrophobic effect.
| S-EPMC2709698 | biostudies-literature
Project description:BackgroundCollagen is a structural protein that provides mechanical stability and defined architectures to skin. In collagen-based skin disorders this stability is lost, either due to mutations in collagens or in the chaperones involved in collagen assembly. This leads to chronic wounds, skin fragility, and blistering. Existing approaches to treat such conditions rely on administration of small molecules to simulate collagen production, like 4-phenylbutyrate (4-PBA) or growth factors like TGF-β. However, these molecules are not specific for collagen synthesis, and result in unsolicited side effects. Hsp47 is a collagen-specific chaperone with a major role in collagen biosynthesis. Expression levels of Hsp47 correlate with collagen deposition. This article explores the stimulation of collagen deposition by exogenously supplied Hsp47 (collagen specific chaperone) to skin cells, including specific collagen subtypes quantification.ResultsHere we quantify the collagen deposition level and the types of deposited collagens after Hsp47 stimulation in different in vitro cultures of cells from human skin tissue (fibroblasts NHDF, keratinocytes HaCat and endothelial cells HDMEC) and mouse fibroblasts (L929 and MEF). We find upregulated deposition of fibrillar collagen subtypes I, III and V after Hsp47 delivery. Network collagen IV deposition was enhanced in HaCat and HDMECs, while fibril-associated collagen XII was not affected by the increased intracellular Hsp47 levels. The deposition levels of fibrillar collagen were cell-dependent i.e. Hsp47-stimulated fibroblasts deposited significantly higher amount of fibrillar collagen than Hsp47-stimulated HaCat and HDMECs.ConclusionsA 3-fold enhancement of collagen deposition was observed in fibroblasts upon repeated dosage of Hsp47 within the first 6 days of culture. Our results provide fundamental understanding towards the idea of using Hsp47 as therapeutic protein to treat collagen disorders.
| S-EPMC7106624 | biostudies-literature
Project description:Molecular dynamics simulations of collagen are used to investigate at the atomistic level the nature of the interprotein interactions that are present within a collagen fibril, and which are responsible for the fibril's thermodynamic stability. Simulations both of a collagen fibril and of a fully solvated tropcollagen are compared in order to study the interactions that arise between the proteins upon the process of fibrillogenesis. The interactions studied include direct interprotein hydrogen bonds, water-mediated interprotein hydrogen bonds, and hydrophobic interactions. The simulations are used to quantify the number of interprotein interactions that form; to study which functional groups contribute most towards the interactions; and to study the spatial distribution of interprotein interactions throughout the fibril's D period. The processes of collagen fibrillogenesis and protein folding are then compared with each other, because these two physical processes share many similarities in concept, and the latter has been more widely studied. Molecular dynamics simulations of a bacteriophage T4 lysozyme protein, both in its native state and in and unfolded state, are used as an illustrative example of a typical protein folding process, for direct comparison with the collagen simulations.
| S-EPMC3605786 | biostudies-literature
Project description:Osteogenesis imperfecta (OI) is a genetic disorder in collagen characterized by mechanically weakened tendon, fragile bones, skeletal deformities, and in severe cases, prenatal death. Although many studies have attempted to associate specific mutation types with phenotypic severity, the molecular and mesoscale mechanisms by which a single point mutation influences the mechanical behavior of tissues at multiple length scales remain unknown. We show by a hierarchy of full atomistic and mesoscale simulation that OI mutations severely compromise the mechanical properties of collagenous tissues at multiple scales, from single molecules to collagen fibrils. Mutations that lead to the most severe OI phenotype correlate with the strongest effects, leading to weakened intermolecular adhesion, increased intermolecular spacing, reduced stiffness, as well as a reduced failure strength of collagen fibrils. We find that these molecular-level changes lead to an alteration of the stress distribution in mutated collagen fibrils, causing the formation of stress concentrations that induce material failure via intermolecular slip. We believe that our findings provide insight into the microscopic mechanisms of this disease and lead to explanations of characteristic OI tissue features such as reduced mechanical strength and a lower cross-link density. Our study explains how single point mutations can control the breakdown of tissue at much larger length scales, a question of great relevance for a broad class of genetic diseases.
| S-EPMC2718154 | biostudies-literature