Molecular Dynamics Investigation on the Effects of Protonation and Lysyl Hydroxylation on Sulfilimine Cross-links in Collagen IV
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PROVIDER: S-EPMC9647856 | biostudies-literature | 2022 Oct
REPOSITORIES: biostudies-literature
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Project description:Collagen IV networks are ancient proteins of basement membranes that underlie epithelia in metazoa from sponge to human. The networks provide structural integrity to tissues and serve as ligands for integrin cell-surface receptors. They are assembled by oligomerization of triple-helical protomers and are covalently crosslinked, a key reinforcement that stabilizes networks. We used Fourier-transform ion cyclotron resonance mass spectrometry and nuclear magnetic resonance spectroscopy to show that a sulfilimine bond (-S=N-) crosslinks hydroxylysine-211 and methionine-93 of adjoining protomers, a bond not previously found in biomolecules. This bond, the nitrogen analog of a sulfoxide, appears to have arisen at the divergence of sponge and cnidaria, an adaptation of the extracellular matrix in response to mechanical stress in metazoan evolution.
| S-EPMC2876822 | biostudies-literature
Project description:Collagen IV is essential component of basement membrane in the tissues. It provides proper cellular structure by the formation of sulfilimine bond (S[double bond, length as m-dash]N) between methionine and lysine or hydroxylysine (cross-links) residues which can be formed with or without post-translational modification. The sulfilimine bond has critical roles in tissue development and human diseases. Peroxidasin, a basement membrane peroxidase, generates reactive halogen species including hypobromous (HOBr) acid and hypochlorous (HOCl) acid which help to form halosulfonium or haloamine. The sulfilamine bond can be formed either by the formation of halosulfonium or by the formation of halomine. The aim of the study is the investigation of the formation of sulfilimine bond and its nature in collagen IV using multi-scale approach that included MD, QM-cluster, systematic series of small models, and NBO analysis. These results suggest that sulfilimine bond can be formed either via brominated/chlorinated halosulfonium or haloamine pathway. The results of systematic series of small model indicate that the formation of sulfilimine complex from halosulfonium happens through the formation of positively charged halosulfonated sulfilimine complex. It also suggests that the formation of sulfilimine complex from haloamine occurs through the formation of positively charged sulfilimine complex where the S and N bond forms and halogen goes off at the same time. Furthermore, the NBO analysis suggest the S and N bond is strongly polarized toward nitrogen in both single protonated and neutral system, N δ- ← S δ+ and also indicate the existence of a coordinate covalent (i.e. dative) bond.
| S-EPMC9306264 | biostudies-literature
Project description:The incubation of lens capsules with glucose in vitro resulted in changes in the mechanical and thermal properties of type-IV collagen consistent with increased cross-linking. Differential scanning calorimetry (d.s.c.) of fresh lens capsules showed two major peaks at melting temperatures Tm 1 and Tm 2 at approx. 54 degrees C and 90 degrees C, which can be attributed to the denaturation of the triple helix and 7S domains respectively. Glycosylation of lens capsules in vitro for 24 weeks caused an increase in Tm 1 from 54 degrees C to 61 degrees C, while non-glycosylated, control incubated capsules increased to a Tm 1 of 57 degrees C. The higher temperature required to denature the type-IV collagen after incubation in vitro suggested increased intermolecular cross-linking. Glycosylated lens capsules were more brittle than fresh samples, breaking at a maximum strain of 36.8 +/- 1.8% compared with 75.6 +/- 6.3% for the fresh samples. The stress at maximum strain (or 'strength') was dramatically reduced from 12.0 to 4.7 N.mm.mg-1 after glycosylation in vitro. The increased constraints within the system leading to loss of strength and increased brittleness suggested not only the presence of more cross-links but a difference in the location of these cross-links compared with the natural lysyl-aldehyde-derived cross-links. The chemical nature of the fluorescent glucose-derived cross-link following glycosylation was determined as pentosidine, at a concentration of 1 pentosidine molecule per 600 collagen molecules after 24 weeks incubation. Pentosidine was also determined in the lens capsules obtained from uncontrolled diabetics at a level of about 1 per 100 collagen molecules. The concentration of these pentosidine cross-links is far too small to account for the observed changes in the thermal and mechanical properties following incubation in vitro, clearly indicating that another as yet undefined, but apparently more important cross-linking mechanism mediated by glucose is taking place.
| S-EPMC1137721 | biostudies-other
Project description:Cardiac fibrosis-mediated heart failure (HF) is one of the major forms of end-stage cardiovascular diseases (CVDs). Cardiac fibrosis is an adaptive response of the myocardium upon any insult/injury. Excessive deposition of collagen molecules in the extracellular matrix (ECM) is the hallmark of fibrosis. This fibrotic response initially protects the myocardium from ventricular rupture. Although in mammals this fibrotic response progresses towards scar-tissue formation leading to HF, some fishes and urodeles have mastered the art of cardiac regeneration following injury-mediated fibrotic response. Zebrafish have a unique capability to regenerate the myocardium after post-amputation injury. Following post-amputation, the ECM of the zebrafish heart undergoes extensive remodeling and deposition of collagen. Being the most abundant protein of ECM, collagen plays important role in the assembly and cell-matrix interactions. However, the mechanism of ECM remodeling is not well understood. Collagen molecules undergo heavy post-translational modifications (PTMs) mainly hydroxylation of proline, lysine, and glycosylation of lysine during biosynthesis. The critical roles of these PTMs are emerging in several diseases, embryonic development, cell behavior regulation, and cell-matrix interactions. The site-specific identification of these collagen PTMs in zebrafish heart ECM is not known. As these highly modified peptides are not amenable to mass spectrometry (MS), the site-specific identification of these collagen PTMs is challenging. Here, we have implemented our in-house proteomics analytical pipeline to analyze two ECM proteomics datasets (PXD011627, PXD010092) of the zebrafish heart during regeneration (post-amputation). We report the first comprehensive site-specific collagen PTM map of zebrafish heart ECM. We have identified a total of 36 collagen chains (19 are reported for the first time here) harboring a total of 95 prolyl-3-hydroxylation, 108 hydroxylysine, 29 galactosyl-hydroxylysine, and 128 glucosylgalactosyl-hydroxylysine sites. Furthermore, we comprehensively map the three chains (COL1A1a, COL1A1b, and COL1A2) of collagen I, the most abundant protein in zebrafish heart ECM. We achieved more than 95% sequence coverage for all the three chains of collagen I. Our analysis also revealed the dynamics of prolyl-3-hydroxylation occupancy oscillations during heart regeneration at these sites. Moreover, quantitative site-specific analysis of lysine-O-glycosylation microheterogeneity during heart regeneration revealed a significant (p < 0.05) elevation of site-specific (K1017) glucosylgalactosyl-hydroxylysine on the col1a1a chain. Taken together, these site-specific PTM maps and the dynamic changes of site-specific collagen PTMs in ECM during heart regeneration will open up new avenues to decode ECM remodeling and may lay the foundation to tinker the cardiac regeneration process with new approaches.
| S-EPMC9245515 | biostudies-literature
Project description:Collagen is a major component of the extracellular matrix and its integrity is essential for connective tissue and organ function. The importance of proteins involved in intracellular collagen post-translational modification, folding and transport was recently highlighted from studies on recessive forms of osteogenesis imperfecta (OI). Here we describe the critical role of SC65 (Synaptonemal Complex 65, P3H4), a leprecan-family member, as part of an endoplasmic reticulum (ER) complex with prolyl 3-hydroxylase 3. This complex affects the activity of lysyl-hydroxylase 1 potentially through interactions with the enzyme and/or cyclophilin B. Loss of Sc65 in the mouse results in instability of this complex, altered collagen lysine hydroxylation and cross-linking leading to connective tissue defects that include low bone mass and skin fragility. This is the first indication of a prolyl-hydroxylase complex in the ER controlling lysyl-hydroxylase activity during collagen synthesis.
| S-EPMC4847768 | biostudies-literature
Project description:Collagen type IV (Col IV) is a basement membrane protein associated with early blood vessel morphogenesis and is essential for blood vessel stability. Defects in vascular Col IV deposition are the basis of heritable disorders, such as small vessel disease, marked by cerebral hemorrhage and drastically shorten lifespan. To date, little is known about how endothelial cells regulate the intracellular transport and selective secretion of Col IV in response to angiogenic cues, leaving a void in our understanding of this critical process. Our aim was to identify trafficking pathways that regulate Col IV deposition during angiogenic blood vessel development. We have identified the GTPase Rab10 as a major regulator of Col IV vesicular trafficking during vascular development using both in vitro imaging and biochemistry as well as in vivo models. Knockdown of Rab10 reduced de novo Col IV secretion in vivo and in vitro. Mechanistically, we determined that Rab10 is an indirect mediator of Col IV secretion, partnering with atypical Rab25 to deliver the enzyme lysyl hydroxylase 3 (LH3) to Col IV-containing vesicles staged for secretion. Loss of Rab10 or Rab25 results in depletion of LH3 from Col IV-containing vesicles and rapid lysosomal degradation of Col IV. Furthermore, we demonstrate that Rab10 is Notch responsive, indicating a novel connection between permissive Notch-based vessel maturation programs and vesicle trafficking. Our results illustrate both a new trafficking-based component in the regulated secretion of Col IV and how this vesicle trafficking program interfaces with Notch signaling to fine-tune basement membrane secretion during blood vessel development.
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Project description:In Duchenne muscular dystrophy (DMD), a lack of functional dystrophin leads to myofiber instability and progressive muscle damage that results in fibrosis. While fibrosis is primarily characterized by an accumulation of extracellular matrix (ECM) components, there are changes in ECM architecture during fibrosis that relate more closely to functional muscle stiffness. One of these architectural changes in dystrophic muscle is collagen cross-linking, which has been shown to increase the passive muscle stiffness in models of fibrosis including the mdx mouse, a model of DMD. We tested whether the intraperitoneal injections of beta-aminopropionitrile (BAPN), an inhibitor of the cross-linking enzyme lysyl oxidase, would reduce collagen cross-linking and passive stiffness in young and adult mdx mice compared to saline-injected controls. We found no significant differences between BAPN treated and saline treated mice in collagen cross-linking and stiffness parameters. However, we observed that while collagen cross-linking and passive stiffness scaled positively in dystrophic muscles, collagen fiber alignment scaled with passive stiffness distinctly between muscles. We also observed that the dystrophic diaphragm showed the most dramatic fibrosis in terms of collagen content, cross-linking, and stiffness. Overall, we show that while BAPN was not effective at reducing collagen cross-linking, the positive association between collagen cross-linking and stiffness in dystrophic muscles still show cross-linking as a viable target for reducing passive muscle stiffness in DMD or other fibrotic muscle conditions.
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Project description:Bruck syndrome (BS) is a disorder characterized by joint flexion contractures and skeletal dysplasia that shows strong clinical overlap with the brittle bone disease osteogenesis imperfecta (OI). BS is caused by biallelic mutations in either the FKBP10 or the PLOD2 gene. PLOD2 encodes the lysyl hydroxylase 2 (LH2) enzyme, which is responsible for the hydroxylation of lysine residues in fibrillar collagen telopeptides. This hydroxylation directs crosslinking of collagen fibrils in the extracellular matrix, which is necessary to provide stability and tensile integrity to the collagen fibrils. To further elucidate the function of LH2 in vertebrate skeletal development, we created a zebrafish model harboring a homozygous plod2 nonsense mutation resulting in reduced telopeptide hydroxylation and crosslinking of bone type I collagen. Adult plod2 mutants present with a shortened body axis and severe skeletal abnormalities with evidence of bone fragility and fractures. The vertebral column of plod2 mutants is short and scoliotic with compressed vertebrae that show excessive bone formation at the vertebral end plates, and increased tissue mineral density in the vertebral centra. The muscle fibers of mutant zebrafish have a reduced diameter near the horizontal myoseptum. The endomysium, a layer of connective tissue ensheathing the individual muscle fibers, is enlarged. Transmission electron microscopy of mutant vertebral bone shows type I collagen fibrils that are less organized with loss of the typical plywood-like structure. In conclusion, plod2 mutant zebrafish show molecular and tissue abnormalities in the musculoskeletal system that are concordant with clinical findings in BS patients. Therefore, the plod2 zebrafish mutant is a promising model for the elucidation of the underlying pathogenetic mechanisms leading to BS and the development of novel therapeutic avenues in this syndrome. © 2016 American Society for Bone and Mineral Research.
| S-EPMC5364950 | biostudies-literature
Project description:Bottom up proteomics, cross-linked amino acid analysis, and cross-linked peptide analysis were performed on eight PDAC tumor samples and paired adjacent non-tumor samples.
2025-08-28 | PXD050833 | Pride
Project description:This paper describes the isolation from reduced collagen of two new amino acids believed to be involved, in their non-reduced form, as intermolecular cross-links stabilizing the collagen fibre. The reduction of intact collagen fibrils with tritiated sodium borohydride was found to stabilize the aldehyde-mediated cross-links to acid hydrolysis and thus allowed their location and isolation from acid hydrolysates on an automatic amino acid analyser. Comparison of the radioactive elution patterns from the autoanalyser of collagen treated in various ways before reduction permitted a preliminary classification of the peaks into cross-link precursors, intramolecular and intermolecular cross-links. The techniques employed to isolate the purified components on a large scale and to identify them structurally are described in detail. Two labile intermolecular cross-links were isolated in their reduced forms, one of which was identified by high-resolution mass spectrometry as N(in)-(5-amino-5-carboxypentyl)hydroxylysine. The structure of this compound was confirmed by chemical synthesis. The cross-link precursor alpha-aminoadipic delta-semialdehyde was isolated in its reduced form, in-hydroxynorleucine, together with its acid degradation product in-chloronorleucine. A relatively stable intermolecular cross-link was isolated and partially characterized by mass spectrometry as an aldol resulting from the reaction of the delta-semialdehyde derived from lysine and hydroxylysine.
| S-EPMC1179041 | biostudies-other