AmyloGraph : a comprehensive database of amyloid-amyloid interactions

Information about the impact of interactions between amyloid proteins on their fibrillization propensity is scattered among many experimental articles and presented in unstructured form. We manually curated information located in almost 200 publications (selected out of 562 initially considered), obtaining details of 883 experimentally studied interactions between 46 amyloid proteins or peptides. We also proposed a novel standardized terminology for the description of amyloid-amyloid interactions, which is included in our database, covering all currently known types of such a cross-talk, including inhibition of fibrillization, cross-seeding and other phenomena. The new approach allows for more specific studies on amyloids and their interactions, by providing very well-defined data. AmyloGraph, an online database presenting information on amyloid-amyloid interactions, is available at (). Its functionalities are also accessible as the R package (). AmyloGraph is the only publicly available repository for experimentally determined amyloid-amyloid interactions

Variability of Amyloid Propensity in Imperfect Repeats of CsgA Protein of Salmonella enterica and Escherichia coli

CsgA is an aggregating protein from bacterial biofilms, representing a class of functional amyloids. Its amyloid propensity is defined by five fragments (R1–R5) of the sequence, representing non-perfect repeats. Gate-keeper amino acid residues, specific to each fragment, define the fragment’s propensity for self-aggregation and aggregating characteristics of the whole protein. We study the self-aggregation and secondary structures of the repeat fragments of Salmonella enterica and Escherichia coli and comparatively analyze their potential effects on these proteins in a bacterial biofilm. Using bioinformatics predictors, ATR-FTIR and FT-Raman spectroscopy techniques, circular dichroism, and transmission electron microscopy, we confirmed self-aggregation of R1, R3, R5 fragments, as previously reported for Escherichia coli, however, with different temporal characteristics for each species. We also observed aggregation propensities of R4 fragment of Salmonella enterica that is different than that of Escherichia coli. Our studies showed that amyloid structures of CsgA repeats are more easily formed and more durable in Salmonella enterica than those in Escherichia coli

Antibody CD133 Biofunctionalization of Ammonium Acryloyldimethyltaurate and Vinylpyrrolidone Co-Polymer-Based Coating of the Vascular Implants

Current vascular stents, such as drug eluting stents (DES), have some serious drawbacks, like in stent restenosis and thrombosis. Therefore, other solutions are sought to overcome these post-implantations complications. These include the strategy of biofunctionalization of the stent surface with antibodies that facilitate adhesion of endothelial cells (ECs) or endothelial progenitor cells (EPCs). Rapid re-endothelialization of the surface minimizes the risk of possible complications. In this study, we proposed ammonium acryloyldimethyltaurate/vinylpyrrolidone co-polymer-based surface (AVC), which was mercaptosilanized in order to expose free thiol groups. The presence of free thiol groups allowed for the covalent attachment of CD133 antibodies by disulfide bridges formation between mercaptosilanized surface and cysteine of the protein molecule thiol groups. Various examinations were performed in order to validate the procedure, including attenuated total reflection–Fourier transform infrared spectroscopy (ATR-FTIR) and Fourier transform Raman spectroscopy (FT-Raman), atomic force microscopy (AFM) and scanning electron microscopy (SEM). By means of ATR-FTIR spectroscopy presence of the CD133 antibody within coating was confirmed. In vitro studies proved good biocompatibility for blood cells without induction of hemolytic response. Thus, proposed biofunctionalized CD133 antibody AVC surface has shown sufficient stability for adapting as cardiovascular implant coating and biocompatibility. According to conducted in vitro studies, the modified surface can be further tested for applications in various biological systems

Applicability of FTIR-ATR Method to Measure Carbonyls in Blood Plasma after Physical and Mental Stress

Introduction. Oxidative stress is a state of imbalance between the production of reactive oxygen species and antioxidant defenses. It results in the oxidation of all cellular elements and, to a large extent, proteins, causing inter alia the formation of carbonyl groups in their structures. The study focused on assessment of changes in the plasma protein-bound carbonyls in police horses after combat training and after rest and the applicability of infrared spectroscopy with a Fourier transform, utilizing the attenuated total reflectance (FTIR-ATR) in detecting plasma protein oxidation. Methods. We evaluated the influence of both the different concentrations of hydrogen peroxide and combat training on protein carbonylation in horse blood plasma. The oxidation of plasma proteins was assessed using a spectrophotometric method based on the carbonyl groups derivatization with 2,4-dinitrophenylhydrazine (DNPH). The measured values were correlated with the carbonyl groups concentrations determined by means of the FTIR-ATR method. Results. The linear correlation between the DNPH and FTIR-ATR methods was shown. The concentration of plasma protein-bound carbonyls significantly deceased in police horses after one-day rest when compared to the values measured directly after the combat training (a drop by 23%, p<0.05 and 29%, p<0.01 measured by DNPH and FTIR-ATR methods, respectively). These results were consistent with the proteins phosphorylation analysis. Conclusion. The FTIR-ATR method may be applied to measure the level of plasma proteins peroxidation

Applicability of FTIR-ATR method to measure carbonyls in blood plasma after physical and mental stress

Introduction. Oxidative stress is a state of imbalance between the production of reactive oxygen species and antioxidant defenses. It results in the oxidation of all cellular elements and, to a large extent, proteins, causing inter alia the formation of carbonyl groups in their structures. The study focused on assessment of changes in the plasma protein-bound carbonyls in police horses after combat training and after rest and the applicability of infrared spectroscopy with a Fourier transform, utilizing the attenuated total reflectance (FTIR-ATR) in detecting plasma protein oxidation. Methods. We evaluated the influence of both the different concentrations of hydrogen peroxide and combat training on protein carbonylation in horse blood plasma. The oxidation of plasma proteins was assessed using a spectrophotometric method based on the carbonyl groups derivatization with 2,4-dinitrophenylhydrazine (DNPH). The measured values were correlated with the carbonyl groups concentrations determined by means of the FTIR-ATR method. Results. The linear correlation between the DNPH and FTIR-ATR methods was shown. The concentration of plasma protein-bound carbonyls significantly deceased in police horses after one-day rest when compared to the values measured directly after the combat training (a drop by 23%, pThis work was financed under the program of the Minister of Science and Higher Education “Strategy of Excellence - University of Research” in 2018-2019 project number 0019/SDU/2018/18 in the amount of PLN 700 000 and partly by the statutory funds of Wrocław University of Science and Technology. This work was created in cooperation with the Ministry of the Interior of the Slovak Republic, Department of Police Kynology and Hippology. Tomasz Walski was supported by the European Union’s Horizon 2020 Research and Innovation Programme, under the Marie Skłodowska-Curie Grant Agreement No 713690, and by the Science Foundation Ireland (SFI) and the European Regional Development Fund (Grant Number 13/RC/2073)

Bioinformatics methods for identification of amyloidogenic peptides show robustness to misannotated training data

Several disorders are related to amyloid aggregation of proteins, for example Alzheimer's or Parkinson's diseases. Amyloid proteins form fibrils of aggregated beta structures. This is preceded by formation of oligomers-the most cytotoxic species. Determining amyloidogenicity is tedious and costly. The most reliable identification of amyloids is obtained with high resolution microscopies, such as electron microscopy or atomic force microscopy (AFM). More frequently, less expensive and faster methods are used, especially infrared (IR) spectroscopy or Thioflavin T staining. Different experimental methods are not always concurrent, especially when amyloid peptides do not readily form fibrils but oligomers. This may lead to peptide misclassification and mislabeling. Several bioinformatics methods have been proposed for in-silico identification of amyloids, many of them based on machine learning. The effectiveness of these methods heavily depends on accurate annotation of the reference training data obtained from in-vitro experiments. We study how robust are bioinformatics methods to weak supervision, encountering imperfect training data. AmyloGram and three other amyloid predictors were applied. The results proved that a certain degree of misannotation in the reference data can be eliminated by the bioinformatics tools, even if they belonged to their training set. The computational results are supported by new experiments with IR and AFM methods

Antiatherosclerotic effects of 1-methylnicotinamide in apolipoprotein E/low-density lipoprotein receptor-deficient mice : a comparison with nicotinic acid

1-Methylnicotinamide (MNA), the major endogenous metabolite of nicotinic acid (NicA), may partially contribute to the vasoprotective properties of NicA. Here we compared the antiatherosclerotic effects of MNA and NicA in apolipoprotein E (ApoE)/ low-density lipoprotein receptor (LDLR)-deficient mice. ApoE/ LDLR2/2 mice were treated with MNA or NicA (100 mg/kg). Plaque size, macrophages, and cholesterol content in the brachiocephalic artery, endothelial function in the aorta, systemic inflammation, platelet activation, as well as the concentration of MNA and its metabolites in plasma and urine were measured. MNA and NicA reduced atherosclerotic plaque area, plaque inflammation, and cholesterol content in the brachiocephalic artery. The antiatherosclerotic actions of MNA and NicA were associated with improved endothelial function, as evidenced by a higher concentration of 6-keto-prostaglandin F1a and nitrite/nitrate in the aortic ring effluent, inhibition of platelets (blunted thromboxane B2 generation), and inhibition of systemic inflammation (lower plasma concentration of serum amyloid P, haptoglobin). NicA treatment resulted in an approximately 2-fold higher concentration of MNA and its metabolites in urine and a 4-fold higher nicotinamide/MNA ratio in plasma, compared with MNA treatment. In summary; MNA displays pronounced antiatherosclerotic action in ApoE/LDLR2/2 mice, an effect associated with an improvement in prostacyclin- and nitric oxide-dependent endothelial function, inhibition of platelet activation, inhibition of inflammatory burden in plaques, and diminished systemic inflammation. Despite substantially higher MNA availability after NicA treatment, compared with an equivalent dose of MNA, the antiatherosclerotic effect of NicA was not stronger. We suggest that detrimental effects of NicA or its metabolites other than MNA may limit beneficial effects of NicA-derived MNA