RESUMEN
Collagen fibrils are the major constituents of the extracellular matrix, which provides structural support to vertebrate connective tissues. It is widely assumed that the superstructure of collagen fibrils is encoded in the primary sequences of the molecular building blocks. However, the interplay between large-scale architecture and small-scale molecular interactions makes the ab initio prediction of collagen structure challenging. Here, we propose a model that allows us to predict the periodic structure of collagen fibers and the axial offset between the molecules, purely on the basis of simple predictive rules for the interaction between amino acid residues. With our model, we identify the sequence-dependent collagen fiber geometries with the lowest free energy and validate the predicted geometries against the available experimental data. We propose a procedure for searching for optimal staggering distances. Finally, we build a classification algorithm and use it to scan 11 data sets of vertebrate fibrillar collagens, and predict the periodicity of the resulting assemblies. We analyzed the experimentally observed variance of the optimal stagger distances across species, and find that these distances, and the resulting fibrillar phenotypes, are evolutionary well preserved. Moreover, we observed that the energy minimum at the optimal stagger distance is broad in all cases, suggesting a further evolutionary adaptation designed to improve the assembly kinetics. Our periodicity predictions are not only in good agreement with the experimental data on collagen molecular staggering for all collagen types analyzed, but also for synthetic peptides. We argue that, with our model, it becomes possible to design tailor-made, periodic collagen structures, thereby enabling the design of novel biomimetic materials based on collagen-mimetic trimers.
Asunto(s)
Materiales Biomiméticos , Colágeno , Materiales Biomiméticos/química , Colágeno/metabolismo , Matriz Extracelular/metabolismo , Colágenos Fibrilares , Péptidos/químicaRESUMEN
Several peptide dual agonists of the human glucagon receptor (GCGR) and the glucagon-like peptide-1 receptor (GLP-1R) are in development for the treatment of type 2 diabetes, obesity and their associated complications. Candidates must have high potency at both receptors, but it is unclear whether the limited experimental data available can be used to train models that accurately predict the activity at both receptors of new peptide variants. Here we use peptide sequence data labelled with in vitro potency at human GCGR and GLP-1R to train several models, including a deep multi-task neural-network model using multiple loss optimization. Model-guided sequence optimization was used to design three groups of peptide variants, with distinct ranges of predicted dual activity. We found that three of the model-designed sequences are potent dual agonists with superior biological activity. With our designs we were able to achieve up to sevenfold potency improvement at both receptors simultaneously compared to the best dual-agonist in the training set.
RESUMEN
Collagen fibrils are central to the molecular organization of the extracellular matrix (ECM) and to defining the cellular microenvironment. Glycation of collagen fibrils is known to impact on cell adhesion and migration in the context of cancer and in model studies, glycation of collagen molecules has been shown to affect the binding of other ECM components to collagen. Here we use TEM to show that ribose-5-phosphate (R5P) glycation of collagen fibrils - potentially important in the microenvironment of actively dividing cells, such as cancer cells - disrupts the longitudinal ordering of the molecules in collagen fibrils and, using KFM and FLiM, that R5P-glycated collagen fibrils have a more negative surface charge than unglycated fibrils. Altered molecular arrangement can be expected to impact on the accessibility of cell adhesion sites and altered fibril surface charge on the integrity of the extracellular matrix structure surrounding glycated collagen fibrils. Both effects are highly relevant for cell adhesion and migration within the tumour microenvironment.
Asunto(s)
Colágeno Tipo I/química , Matriz Extracelular/química , Ribosamonofosfatos/química , Animales , Colágeno Tipo I/metabolismo , Matriz Extracelular/metabolismo , Glicosilación , Humanos , Ribosamonofosfatos/metabolismoRESUMEN
Arterial hypertension is considered to be an inflammatory condition with low intensity. Therefore, an elevated concentration of inflammatory cytokines can be expected in patients with systemic arterial hypertension, including tumor necrosis factor (TNF).The study included a group of 96 persons aged 18 to 65 years: 76 patients with primary arterial hypertension and 20 healthy individuals (control group). Blood pressure was measured in all individuals using the office and ambulatory blood pressure monitoring (ABPM) measurement, blood was collected for laboratory tests [tumor necrosis factor (TNF), tumor necrosis factor receptor 1 (TNFR1)], and 24-hour urine collection was performed in which albuminuria and TNF concentration were assessed. Moreover, assessment of the intima-media thickness (IMT) in ultrasonography and left ventricular mass index (LVMI) in echocardiography were carried out.Statistically elevated TNF concentration in the blood serum (Pâ=â.0001) and in the 24-hour urine collection (Pâ=â.0087) was determined in patients with hypertension in comparison with the control group. The TNF and TNFR1 concentration in the serum and TNF in the 24-hour urine in the group of patients with arterial hypertension and organ damages and without such complications did not differ statistically significantly.We observed a positive and statistically significant correlation between TNFR1 concentration in the serum and TNF urine excretion in patients with hypertension (râ=â0.369, Pâ<â.05)Patients with arterial hypertension are characterized by higher TNF concentrations in blood serum and higher TNF excretion in 24-hour urine than healthy persons.TNF and TNFR1 concentration in blood serum and TNF excretion in 24-hour urine in patients with early organ damages due to arterial hypertension do not differ significantly from those parameters in patients with arterial hypertension without organ complications.There is a positive correlation between TNFR1 concentration in the serum and TNF urine excretion in patients with hypertension.
Asunto(s)
Albuminuria/orina , Hipertensión/sangre , Hipertensión/orina , Factor de Necrosis Tumoral alfa/sangre , Factor de Necrosis Tumoral alfa/orina , Adolescente , Adulto , Anciano , Albuminuria/etiología , Monitoreo Ambulatorio de la Presión Arterial , Grosor Intima-Media Carotídeo , Estudios de Casos y Controles , Ecocardiografía , Femenino , Humanos , Masculino , Persona de Mediana Edad , Receptores Tipo I de Factores de Necrosis Tumoral/sangre , Receptores Tipo I de Factores de Necrosis Tumoral/orina , Adulto JovenRESUMEN
Sensitivity enhancement by isotope enrichment and DNP NMR enables detection of minor but biologically relevant species in native intact bone, including nucleic acids, choline from phospholipid headgroups, and histidinyl and hydroxylysyl groups. Labelled matrix from the aggressive osteosarcoma K7M2 cell line confirms the assignments of nucleic acid signals arising from purine, pyrimidine, ribose, and deoxyribose species. Detection of these species is an important and necessary step in elucidating the atomic level structural basis of their functions in intact tissue.
RESUMEN
Biomineralization of the extracellular matrix is an essential, regulated process. Inappropriate mineralization of bone and the vasculature has devastating effects on patient health, yet an integrated understanding of the chemical and cell biological processes that lead to mineral nucleation remains elusive. Here, we report that biomineralization of bone and the vasculature is associated with extracellular poly(ADP-ribose) synthesized by poly(ADP-ribose) polymerases in response to oxidative and/or DNA damage. We use ultrastructural methods to show poly(ADP-ribose) can form both calcified spherical particles, reminiscent of those found in vascular calcification, and biomimetically calcified collagen fibrils similar to bone. Importantly, inhibition of poly(ADP-ribose) biosynthesis in vitro and in vivo inhibits biomineralization, suggesting a therapeutic route for the treatment of vascular calcifications. We conclude that poly(ADP-ribose) plays a central chemical role in both pathological and physiological extracellular matrix calcification.
Asunto(s)
Biomineralización , Daño del ADN , Poli Adenosina Difosfato Ribosa/metabolismo , Calcificación Vascular/metabolismo , Adolescente , Adulto , Anciano , Animales , Vasos Sanguíneos/metabolismo , Vasos Sanguíneos/patología , Bovinos , Línea Celular , Células Cultivadas , Colágeno/metabolismo , Matriz Extracelular/metabolismo , Femenino , Humanos , Masculino , Ratones , Persona de Mediana Edad , Osteoblastos/metabolismo , Osteoblastos/patología , Estrés Oxidativo , Ratas , Ratas Wistar , OvinosRESUMEN
Fibrillar collagens have mechanical and biological roles, providing tissues with both tensile strength and cell binding sites which allow molecular interactions with cell-surface receptors such as integrins. A key question is: how do collagens allow tissue flexibility whilst maintaining well-defined ligand binding sites? Here we show that proline residues in collagen glycine-proline-hydroxyproline (Gly-Pro-Hyp) triplets provide local conformational flexibility, which in turn confers well-defined, low energy molecular compression-extension and bending, by employing two-dimensional 13C-13C correlation NMR spectroscopy on 13C-labelled intact ex vivo bone and in vitro osteoblast extracellular matrix. We also find that the positions of Gly-Pro-Hyp triplets are highly conserved between animal species, and are spatially clustered in the currently-accepted model of molecular ordering in collagen type I fibrils. We propose that the Gly-Pro-Hyp triplets in fibrillar collagens provide fibril "expansion joints" to maintain molecular ordering within the fibril, thereby preserving the structural integrity of ligand binding sites.