SwePub - sökning: WFRF:(Bernfur Katja)

Numrering	Referens	Omslagsbild	Hitta
1.	Ahlgren, Eva Christina, et al. (författare) Iron-induced oligomerization of human FXN81-210 and bacterial CyaY frataxin and the effect of iron chelators 2017 Ingår i: PLoS ONE. - : Public Library of Science (PLoS). - 1932-6203. ; 12:12 Tidskriftsartikel (refereegranskat)abstract Patients suffering from the progressive neurodegenerative disease Friedreich’s ataxia have reduced expression levels of the protein frataxin. Three major isoforms of human frataxin have been identified, FXN42-210, FXN56-210 and FXN81-210, of which FXN81-210 is considered to be the mature form. Both long forms, FXN42-210 and FXN56-210, have been shown to spontaneously form oligomeric particles stabilized by the extended N-terminal sequence. The short variant FXN81-210, on other hand, has only been observed in the monomeric state. However, a highly homologous E. coli frataxin CyaY, which also lacks an N-terminal extension, has been shown to oligomerize in the presence of iron. To explore the mechanisms of stabilization of short variant frataxin oligomers we compare here the effect of iron on the oligomerization of CyaY and FXN81-210. Using dynamic light scattering, small-angle X-ray scattering, electron microscopy (EM) and cross linking mass spectrometry (MS), we show that at aerobic conditions in the presence of iron both FXN81-210 and CyaY form oligomers. However, while CyaY oligomers are stable over time, FXN81-210 oligomers are unstable and dissociate into monomers after about 24 h. EM and MS studies suggest that within the oligomers FXN81-210 and CyaY monomers are packed in a head-to-tail fashion in ring-shaped structures with potential iron-binding sites located at the interface between monomers. The higher stability of CyaY oligomers can be explained by a higher number of acidic residues at the interface between monomers, which may result in a more stable iron binding. We also show that CyaY oligomers may be dissociated by ferric iron chelators deferiprone and DFO, as well as by the ferrous iron chelator BIPY. Surprisingly, deferiprone and DFO stimulate FXN81-210 oligomerization, while BIPY does not show any effect on oligomerization in this case. The results suggest that FXN81-210 oligomerization is primarily driven by ferric iron, while both ferric and ferrous iron participate in CyaY oligomer stabilization. Analysis of the amino acid sequences of bacterial and eukaryotic frataxins suggests that variations in the position of the acidic residues in helix 1, β-strand 1 and the loop between them may control the mode of frataxin oligomerization.
2.	Alexandersson, Erik, et al. (författare) Plasma membrane proteomics 2006 Ingår i: Plant Proteomics. - Berlin, Heidelberg : Springer Berlin Heidelberg. - 9783540726166 - 9783540726173 ; , s. 186-206 Bokkapitel (refereegranskat)abstract Proteins residing in the plasma membrane have key functions in transport, signal transduction, vesicle trafficking and many other important processes. To better understand these processes it is necessary to reveal the identity of plasma membrane proteins and to monitor modifications and regulation of their expression. This chapter is an overview of the methods used in plant plasma membrane proteomic studies and the results obtained so far. It focuses on studies using mass spectrometry for identification and includes aspects of plasma membrane fractionation, extraction and washing treatments, assessment of purity, separation methods for plasma membrane proteins and choice of techniques for protein cleavage. Finally, the results of plasma membrane proteomic studies are compared and problems with contaminating proteins are discussed.
3.	Alexandersson, Erik, et al. (författare) Purification and proteomic analysis of plant plasma membranes. 2008 Ingår i: Methods in Molecular Biology. - Totowa, NJ : Humana Press. - 1940-6029. ; 432, s. 161-173 Tidskriftsartikel (refereegranskat)abstract All techniques needed for proteomic analyses of plant plasma membranes are described in detail, from isolation of plasma membranes to protein identification by mass spectrometry (MS). Plasma membranes are isolated by aqueous two-phase partitioning yielding vesicles with a cytoplasmic side-in orientation and a purity of about 95%. These vesicles are turned inside-out by treatment with Brij 58, which removes soluble contaminating proteins enclosed in the vesicles as well as loosely attached proteins. The final plasma membrane preparation thus retains all integral proteins and many peripheral proteins. Proteins are separated by one-dimensional sodium dodecyl sulphate-polyacrylamide gel electrophoresis (SDS-PAGE), and protein bands are excised and digested with trypsin. Peptides in tryptic digests are separated by nanoflow liquid chromatography and either fed directly into an ESI-MS or spotted onto matrix-assisted laser desorption ionization (MALDI) plates for analysis with MALDI-MS. Finally, data processing and database searching are used for protein identification to define a plasma membrane proteome.
4.	Alkhalili, Rawana N., et al. (författare) Antimicrobial protein candidates from the thermophilic Geobacillus sp. Strain ZGt-1 : Production, proteomics, and bioinformatics analysis 2016 Ingår i: International Journal of Molecular Sciences. - : MDPI AG. - 1661-6596 .- 1422-0067. ; 17:8 Tidskriftsartikel (refereegranskat)abstract A thermophilic bacterial strain, Geobacillus sp. ZGt-1, isolated from Zara hot spring in Jordan, was capable of inhibiting the growth of the thermophilic G. stearothermophilus and the mesophilic Bacillus subtilis and Salmonella typhimurium on a solid cultivation medium. Antibacterial activity was not observed when ZGt-1 was cultivated in a liquid medium; however, immobilization of the cells in agar beads that were subjected to sequential batch cultivation in the liquid medium at 60 °C showed increasing antibacterial activity up to 14 cycles. The antibacterial activity was lost on protease treatment of the culture supernatant. Concentration of the protein fraction by ammonium sulphate precipitation followed by denaturing polyacrylamide gel electrophoresis separation and analysis of the gel for antibacterial activity against G. stearothermophilus showed a distinct inhibition zone in 15-20 kDa range, suggesting that the active molecule(s) are resistant to denaturation by SDS. Mass spectrometric analysis of the protein bands around the active region resulted in identification of 22 proteins with molecular weight in the range of interest, three of which were new and are here proposed as potential antimicrobial protein candidates by in silico analysis of their amino acid sequences. Mass spectrometric analysis also indicated the presence of partial sequences of antimicrobial enzymes, amidase, and DD-carboxypeptidase.
5.	Bernfur, Katja (författare) Plant Membrane Proteomics - using isolated membranes and proteins to compare and quantify subproteomes 2014 Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract Membrane proteins are particularly important to characterize, since they are involved in cellular processes of utmost importance in control and regulation of cells, such as transport across membranes, signal transduction and photosynthesis. The word "proteome" means all proteins expressed by a e.g. a cell or a tissue, at a certain time point and "proteomics" is the large scale study of these proteins. This thesis concerns mass spectrometry based quantitative proteomics of isolated plant membranes and proteins to gain more insights into the subproteomes. An important prerequisite for the investigations of plant plasma membrane (PM) subproteomes is the isolation of pure membranes, which we obtained by using aqueous two-phase partitioning (paper I). By using highly pure membranes, we were able to identify and quantify tissue specific isoforms in both poplar (Paper II) and Arabidopsis (Paper III). A large number of proteins, unique to the wood-forming xylem PM of poplar, were identified. We applied a quantitative approach using metabolic labeling and mass spectrometric determination of 14N/15N-ratios to determine the distribution differences for e.g. PM aquaporin isofoms, between leaf and root tissue in Arabidopsis (Paper III). Focus was then shifted to the Arabidopsis subproteome and the chloroplast-localized small heat shock protein, Hsp21 (Paper IV). This protein is plays a crucial role for plant survival during periods of stress, by interacting with and preventing stress-sensitive proteins from unfolding. Using our metabolic labeling approach and 14N/15N-ratios, we could detect and quantify the translocation of Hsp21 from the soluble stromal phase to the thylakoid membrane in heat-stressed plants, which was quite unique for Hsp21 compared to other thylakoid membrane proteins (Paper V). Quantitative proteomics using metabolic labeling and 14N/15N-ratios has proven to be a robust and useful tool to investigate differences in plant subproteomes and to investigate the location, abundance and behavior of membrane proteins both in vivo and in vitro.
6.	Bernfur, Katja, et al. (författare) Relative abundance of integral plasma membrane proteins in Arabidopsis leaf and root tissue determined by metabolic labeling and mass spectrometry. 2013 Ingår i: PLoS ONE. - : Public Library of Science (PLoS). - 1932-6203. ; 8:8 Tidskriftsartikel (refereegranskat)abstract Metabolic labeling of proteins with a stable isotope ((15)N) in intact Arabidopsis plants was used for accurate determination by mass spectrometry of differences in protein abundance between plasma membranes isolated from leaves and roots. In total, 703 proteins were identified, of which 188 were predicted to be integral membrane proteins. Major classes were transporters, receptors, proteins involved in membrane trafficking and cell wall-related proteins. Forty-one of the integral proteins, including nine of the 13 isoforms of the PIP (plasma membrane intrinsic protein) aquaporin subfamily, could be identified by peptides unique to these proteins, which made it possible to determine their relative abundance in leaf and root tissue. In addition, peptides shared between isoforms gave information on the proportions of these isoforms. A comparison between our data for protein levels and corresponding data for mRNA levels in the widely used database Genevestigator showed an agreement for only about two thirds of the proteins. By contrast, localization data available in the literature for 21 of the 41 proteins show a much better agreement with our data, in particular data based on immunostaining of proteins and GUS-staining of promoter activity. Thus, although mRNA levels may provide a useful approximation for protein levels, detection and quantification of isoform-specific peptides by proteomics should generate the most reliable data for the proteome.
7.	Bernfur, Katja, et al. (författare) The chloroplast-localized small heat shock protein Hsp21 associates with the thylakoid membranes in heat-stressed plants 2017 Ingår i: Protein Science. - : Wiley. - 0961-8368. ; 26:9, s. 1773-1784 Tidskriftsartikel (refereegranskat)abstract The small heat shock protein (sHsp) chaperones are crucial for cell survival and can prevent aggregation of client proteins that partially unfold under destabilizing conditions. Most investigations on the chaperone activity of sHsps are based on a limited set of thermosensitive model substrate client proteins since the endogenous targets are often not known. There is a high diversity among sHsps with a single conserved β-sandwich fold domain defining the family, the α-crystallin domain, whereas the N-terminal and C-terminal regions are highly variable in length and sequence among various sHsps and conserved only within orthologues. The endogenous targets are probably also varying among various sHsps, cellular compartments, cell type and organism. Here we have investigated Hsp21, a non-metazoan sHsp expressed in the chloroplasts in green plants which experience huge environmental fluctuations not least in temperature. We describe how Hsp21 can also interact with the chloroplast thylakoid membranes, both when isolated thylakoid membranes are incubated with Hsp21 protein and when plants are heat-stressed. The amount of Hsp21 associated with the thylakoid membranes was precisely determined by quantitative mass spectrometry after metabolic 15N-isotope labeling of either recombinantly expressed and purified Hsp21 protein or intact Arabidopsis thaliana plants. We found that Hsp21 is among few proteins that become associated with the thylakoid membranes in heat-stressed plants, and that approximately two thirds of the pool of chloroplast Hsp21 is affected. We conclude that for a complete picture of the role of sHsps in plant stress resistance also their association with the membranes should be considered.
8.	Cukalevski, Risto, et al. (författare) The A beta 40 and A beta 42 peptides self-assemble into separate homomolecular fibrils in binary mixtures but cross-react during primary nucleation 2015 Ingår i: Chemical Science. - : Royal Society of Chemistry (RSC). - 2041-6539 .- 2041-6520. ; 6:7, s. 4215-4233 Tidskriftsartikel (refereegranskat)abstract The assembly of proteins into amyloid fibrils, a phenomenon central to several currently incurable human diseases, is a process of high specificity that commonly tolerates only a low level of sequence mismatch in the component polypeptides. However, in many cases aggregation-prone polypeptides exist as mixtures with variations in sequence length or post-translational modifications; in particular amyloid beta (A beta) peptides of variable length coexist in the central nervous system and possess a propensity to aggregate in Alzheimer's disease and related dementias. Here we have probed the co-aggregation and cross-seeding behavior of the two principal forms of A beta, A beta 40 and A beta 42 that differ by two hydrophobic residues at the C-terminus. We find, using isotope-labeling, mass spectrometry and electron microscopy that they separate preferentially into homomolecular pure A beta 42 and A beta 40 structures during fibril formation from mixed solutions of both peptides. Although mixed fibrils are not formed, the kinetics of amyloid formation of one peptide is affected by the presence of the other form. In particular monomeric A beta 42 accelerates strongly the aggregation of A beta 40 in a concentration-dependent manner. Whereas the aggregation of each peptide is catalyzed by low concentrations of preformed fibrils of the same peptide, we observe a comparably insignificant effect when A beta 42 fibrils are added to A beta 40 monomer or vice versa. Therefore we conclude that fibril-catalysed nucleus formation and elongation are highly sequence specific events but A beta 40 and A beta 42 interact during primary nucleation. These results provide a molecular level description of homomolecular and heteromolecular aggregation steps in mixtures of polypeptide sequence variants.
9.	Domingo-Espín, Joan, et al. (författare) Site-specific glycations of apolipoprotein A-I lead to differentiated functional effects on lipid-binding and on glucose metabolism 2018 Ingår i: Biochimica et Biophysica Acta - Molecular Basis of Disease. - : Elsevier BV. - 0925-4439. ; 1864:9, s. 2822-2834 Tidskriftsartikel (refereegranskat)abstract Prolonged hyperglycemia in poorly controlled diabetes leads to an increase in reactive glucose metabolites that covalently modify proteins by non-enzymatic glycation reactions. Apolipoprotein A-I (apoA-I) of high-density lipoprotein (HDL) is one of the proteins that becomes glycated in hyperglycemia. The impact of glycation on apoA-I protein structure and function in lipid and glucose metabolism were investigated. ApoA-I was chemically glycated by two different glucose metabolites (methylglyoxal and glycolaldehyde). Synchrotron radiation and conventional circular dichroism spectroscopy were used to study apoA-I structure and stability. The ability to bind lipids was measured by lipid-clearance assay and native gel analysis, and cholesterol efflux was measured by using lipid-laden J774 macrophages. Diet induced obese mice with established insulin resistance, L6 rat and C2C12 mouse myocytes, as well as INS-1E rat insulinoma cells, were used to determine in vivo and in vitro glucose uptake and insulin secretion. Site-specific, covalent modifications of apoA-I (lysines or arginines) led to altered protein structure, reduced lipid binding capability and a reduced ability to catalyze cholesterol efflux from macrophages, partly in a modification-specific manner. The stimulatory effects of apoA-I on the in vivo glucose clearance were negatively affected when apoA-I was modified with methylglyoxal, but not with glycolaldehyde. The in vitro data showed that both glucose uptake in muscle cells and insulin secretion from beta cells were affected. Taken together, glycation modifications impair the apoA-I protein functionality in lipid and glucose metabolism, which is expected to have implications for diabetes patients with poorly controlled blood glucose.
10.	Frankel, Rebecca, et al. (författare) Purification and HDL-like particle formation of apolipoprotein A-I after co-expression with the EDDIE mutant of Npro autoprotease 2021 Ingår i: Protein Expression and Purification. - : Elsevier BV. - 1046-5928. ; 187 Tidskriftsartikel (refereegranskat)abstract Apolipoprotein A-I (ApoA-I) is the major protein constituent of high-density lipoprotein particles, and as such is involved in cholesterol transport and activation of LCAT (the lecithin:cholesterol acyltransferase). It may also form amyloidal deposits in the body, showing the multifaceted interactions of ApoA-I. In order to facilitate the study of ApoA-I in various systems, we have developed a protocol based on recombinant expression in E. coli. ApoA-I is protected from degradation by driving its expression to inclusion bodies using a tag: the EDDIE mutant of Npro autoprotease from classical swine fever virus. Upon refolding, EDDIE will cleave itself off from the target protein. The result is a tag-free ApoA-I, with its N-terminus intact. ApoA-I was then purified using a five-step procedure composed of anion exchange chromatography, immobilized metal ion affinity chromatography, hydrophobic interaction chromatography, boiling and size exclusion chromatography. This led to protein of high purity as confirmed with SDS-PAGE and mass spectrometry. The purified ApoA-I formed discoidal objects in the presence of zwitterionic phospholipid DMPC, showing its retained function of interacting with lipids. The protocol was also tested by expression and purification of two ApoA-I mutants, both of which could be purified in the same manner as the wildtype, showing the robustness of the protocol.
11.	Fridolf, Simon, et al. (författare) Ganglioside GM3 stimulates lipid-protein co-assembly in α-synuclein amyloid formation 2023 Ingår i: Biophysical Chemistry. - : Elsevier BV. - 0301-4622. ; 293 Tidskriftsartikel (refereegranskat)abstract Parkinson's disease is characterized by the aggregation of the presynaptic protein α-synuclein (αSyn), and its co-assembly with lipids and other cellular matter in the brain. Here we investigated lipid-protein co-assembly in a system composed of αSyn and model membranes containing the glycolipid ganglioside GM3. We quantified the uptake of lipids into the co-assembled aggregates and investigated how lipid molecular dynamics is altered by being present in the co-assemblies using solution 1H- and solid-state 13C NMR spectroscopy. Aggregate morphology was studied using cryo-TEM. The overall lipid uptake in the co-assembled aggregates was found to increase with the molar ratio of GM3 in the vesicles. The lipids present in the co-assembled aggregates have reduced acyl chain and headgroup dynamics compared to the protein-free bilayer system. These findings may improve our understanding of how different types of lipids can influence the composition of αSyn aggregates, which may have consequences for amyloid formation in vivo.
12.	Gunnarsson, Stefán B., et al. (författare) Analysis of complexes formed by small gold nanoparticles in low concentration in cell culture media 2019 Ingår i: PLoS ONE. - : Public Library of Science (PLoS). - 1932-6203. ; 14:6 Tidskriftsartikel (refereegranskat)abstract New nanomaterials are constantly developed with applications in everything from cosmetics to high tech electronics. Assessing their biological impact has been done by analysis of their adsorbed protein corona, in vitro cell assays, and larger scale ecotoxicological studies. This has proved to be a huge challenge due to the wide range of available nanomaterials and their unpredictable behaviour in different environments. Furthermore, the enormous number of experimental variables make comparisons difficult. Concentration is one of these variables and can vary greatly depending on the aim of the study. When analysing the protein corona, concentrations are often higher than in cell assays. Using a combination of complementary techniques, we have characterised 20 nm gold nanoparticles in a concentration level commonly used in cell studies. We compare their behaviour in a commonly used, protein rich medium and one protein poor medium over 24 hours. Under these conditions, the NPs were stable in protein rich environment but underwent gradual aggregation in protein poor medium. We characterise the biomolecular corona in both media. In protein poor medium, we can describe the often overlooked aggregation. The aggregates' morphology is confirmed by cryo-TEM. Finally, in the protein poor medium, by infrared spectroscopy, we have identified the amino acid arginine in the biomolecular corona which drives the aggregation.
13.	Gunnarsson, Stefán B., et al. (författare) Analysis of nanoparticle biomolecule complexes 2018 Ingår i: Nanoscale. - : Royal Society of Chemistry (RSC). - 2040-3364 .- 2040-3372. ; 10:9, s. 4246-4257 Tidskriftsartikel (refereegranskat)abstract Nanoparticles exposed to biological fluids adsorb biomolecules on their surface forming a biomolecular corona. This corona determines, on a molecular level, the interactions and impact the newly formed complex has on cells and organisms. The corona formation as well as the physiological and toxicological relevance are commonly investigated. However, an acknowledged but rarely addressed problem in many fields of nanobiotechnology is aggregation and broadened size distribution of nanoparticles following their interactions with the molecules of biological fluids. In blood serum, TiO2 nanoparticles form complexes with a size distribution from 30 nm to more than 500 nm. In this study we have separated these complexes, with good resolution, using preparative centrifugation in a sucrose gradient. Two main apparent size populations were obtained, a fast sedimenting population of complexes that formed a pellet in the preparative centrifugation tube, and a slow sedimenting complex population still suspended in the gradient after centrifugation. Concentration and surface area dependent differences are found in the biomolecular corona between the slow and fast sedimenting fractions. There are more immunoglobulins, lipid binding proteins, and lipid-rich complexes at higher serum concentrations. Sedimentation rate and the biomolecular corona are important factors for evaluating any experiment including nanoparticle exposure. Our results show that traditional description of nanoparticles in biological fluids is an oversimplification and that more thorough characterisations are needed.
14.	Gunnarsson, Stefán Bragi, et al. (författare) Dual topography of laminin corona on gallium arsenide nanowires 2020 Ingår i: Biointerphases. - : American Vacuum Society. - 1934-8630 .- 1559-4106. ; 15:5 Tidskriftsartikel (refereegranskat)abstract Nanowires (NWs) are novel nanomaterials with applications in everything from medical implants to solar cells. With increasing number of applications, it is increasingly likely that organisms are exposed to these materials either intentionally or by accident. It is, therefore, important to study their interactions with biological systems and biomolecules. Upon exposure to biological fluids, nanostructure surfaces are quickly covered by a biomolecule corona. The composition of the corona determines the nanostructure's biological fate. Furthermore, upon adsorption, the protein structure can be affected. In order to study the corona morphology, we used two model proteins, laminin of the extracellular matrix and the immune system enzyme myeloperoxidase. We image the protein corona directly by cryo-TEM and enhance resolution by labeling the corona with activated gold nanoparticles. Three-dimensional imaging of the protein corona further increases the resolution and reveals irregularities in corona topography. By doing so, we identified bimodal distribution of spacing between gold nanoparticles and the NW surface for laminin corona at 58 and 85 nm distance from the NWs' surface. The dual topography of the corona is adding a new complexity of the protein corona surface and its interactions with the surrounding biology.
15.	Haikal, Caroline, et al. (författare) The bacterial amyloids phenol soluble modulins from staphylococcus aureus catalyze alpha-synuclein aggregation 2021 Ingår i: International Journal of Molecular Sciences. - : MDPI AG. - 1661-6596 .- 1422-0067. ; 22:21 Tidskriftsartikel (refereegranskat)abstract Aggregated α-synuclein (α-syn) is the main constituent of Lewy bodies, which are a pathological hallmark of Parkinson’s disease (PD). Environmental factors are thought to be potential triggers capable of initiating the aggregation of the otherwise monomeric α-syn. Braak’s seminal work redirected attention to the intestine and recent reports of dysbiosis have highlighted the potential causative role of the microbiome in the initiation of pathology of PD. Staphylococcus aureus is a bacterium carried by 30–70% of the general population. It has been shown to produce functional amy-loids, called phenol soluble modulins (PSMαs). Here, we studied the kinetics of α-syn aggregation under quiescent conditions in the presence or absence of four different PSMα peptides and observed a remarkable shortening of the lag phase in their presence. Whereas pure α-syn monomer did not aggregate up to 450 h after initiation of the experiment in neither neutral nor mildly acidic buffer, the addition of different PSMα peptides resulted in an almost immediate increase in the Thioflavin T (ThT) fluorescence. Despite similar peptide sequences, the different PSMα peptides displayed distinct effects on the kinetics of α-syn aggregation. Kinetic analyses of the data suggest that all four peptides catalyze α-syn aggregation through heterogeneous primary nucleation. The immunogold electron microscopic analyses showed that the aggregates were fibrillar and composed of α-syn. In addition of the co-aggregated materials to a cell model expressing the A53T α-syn variant fused to GFP was found to catalyze α-syn aggregation and phosphorylation in the cells. Our results provide evidence of a potential trigger of synucleinopathies and could have implications for the prevention of the diseases.
16.	Iacobucci, C., et al. (författare) First Community-Wide, Comparative Cross-Linking Mass Spectrometry Study 2019 Ingår i: Analytical Chemistry. - : American Chemical Society (ACS). - 1520-6882 .- 0003-2700. ; 91:11, s. 6953-6961 Tidskriftsartikel (refereegranskat)abstract The number of publications in the field of chemical cross-linking combined with mass spectrometry (XL-MS) to derive constraints for protein three-dimensional structure modeling and to probe protein-protein interactions has increased during the last years. As the technique is now becoming routine for in vitro and in vivo applications in proteomics and structural biology there is a pressing need to define protocols as well as data analysis and reporting formats. Such consensus formats should become accepted in the field and be shown to lead to reproducible results. This first, community-based harmonization study on XL-MS is based on the results of 32 groups participating worldwide. The aim of this paper is to summarize the status quo of XL-MS and to compare and evaluate existing cross-linking strategies. Our study therefore builds the framework for establishing best practice guidelines to conduct cross-linking experiments, perform data analysis, and define reporting formats with the ultimate goal of assisting scientists to generate accurate and reproducible XL-MS results. © 2019 American Chemical Society.
17.	Kelpsiene, Egle, et al. (författare) Protein binding on acutely toxic and non-toxic polystyrene nanoparticles during filtration by Daphnia magna 2022 Ingår i: Environmental Science: Nano. - : Royal Society of Chemistry (RSC). - 2051-8153 .- 2051-8161. Tidskriftsartikel (refereegranskat)abstract Nanomaterials can adsorb biomolecules to their surface and form a protein corona. Here we investigated the protein profile bound to different sizes of aminated and carboxylated polystyrene (PS) nanoparticles after passing through the digestive tract of the freshwater zooplankter Daphnia magna. We found that acutely toxic aminated 53 nm PS nanoparticles bind a different set of proteins compared to other non-toxic PS nanoparticles. The aminated PS nanoparticles bind a higher number of proteins, which are smaller and more acidic, compared to the proteins which bind to the PS nanoparticles that are non-toxic in acute toxicity tests. The proteins bound to toxic nanoparticles can be divided into two groups. One group of proteins which function is related to the digestive system, whereas the other group of proteins can be related to the epithelium, intracellular structures and processes. Finally, we observed that not only proteins bind to the surfaces of the nanoparticles. Triglycerides effectively bind to 200 nm carboxylated PS nanoparticles but not to the other tested nanoparticles. These results provide information about the composition of the corona formed on surfaces of nanoparticles after a short-term incubation with D. magna and give insights to what underlies the acute toxicity caused by nanoparticles.
18.	Kelpsiene, Egle, et al. (författare) The effect of natural biomolecules on yttrium oxide nanoparticles from a Daphnia magna survival rate perspective 2023 Ingår i: Nanotoxicology. - : Taylor and Francis Ltd.. - 1743-5390 .- 1743-5404. ; 17:4, s. 385-399 Tidskriftsartikel (refereegranskat)abstract The attention to rare earth oxide nanoparticles (NPs), including yttrium oxide (Y2O3), has increased in many fields due to their unique structural characteristics and functional properties. The aim of our study was to investigate the mechanisms by which bio-corona formation on Y2O3 NPs affects their environmental fate and toxicity. The Y2O3 NPs induced toxicity to freshwater filter feeder Daphnia magna at particle concentrations of 1 and 10 mg/L, regardless of particle size. Interactions between naturally excreted biomolecules (e.g. protein, lipids, and polysaccharides) derived from D. magna, and the Y2O3 NPs (30–45 nm) resulted in the formation of an eco-corona, which reduced their toxic effects toward D. magna at a particle concentration of 10 mg/L. No effects were observed at lower concentrations or for the other particle sizes investigated. Copper-zinc (Cu-Zn) superoxide dismutase, apolipophorins, and vitellogenin-1 proteins proved to be the most prominent proteins of the adsorbed corona, and possibly a reason for the reduced toxicity of the 30–45 nm Y2O3 NPs toward D. magna.
19.	Lambert, Wietske, et al. (författare) Probing the transient interaction between the small heat-shock protein Hsp21 and a model substrate protein using crosslinking mass spectrometry. 2012 Ingår i: Cell Stress & Chaperones. - : Springer Science and Business Media LLC. - 1466-1268 .- 1355-8145. Tidskriftsartikel (refereegranskat)abstract Small heat-shock protein chaperones are important players in the protein quality control system of the cell, because they can immediately respond to partially unfolded proteins, thereby protecting the cell from harmful aggregates. The small heat-shock proteins can form large polydisperse oligomers that are exceptionally dynamic, which is implicated in their function of protecting substrate proteins from aggregation. Yet the mechanism of substrate recognition remains poorly understood, and little is known about what parts of the small heat-shock proteins interact with substrates and what parts of a partially unfolded substrate protein interact with the small heat-shock proteins. The transient nature of the interactions that prevent substrate aggregation rationalize probing this interaction by crosslinking mass spectrometry. Here, we used a workflow with lysine-specific crosslinking and offline nano-liquid chromatography matrix-assisted laser desorption/ionization tandem time-of-flight mass spectrometry to explore the interaction between the plant small heat-shock protein Hsp21 and a thermosensitive model substrate protein, malate dehydrogenase. The identified crosslinks point at an interaction between the disordered N-terminal region of Hsp21 and the C-terminal presumably unfolding part of the substrate protein.
20.	Lattanzi, Veronica, et al. (författare) Solubility of Aβ40 peptide 2021 Ingår i: JCIS Open. - : Elsevier BV. - 2666-934X. ; 4 Tidskriftsartikel (refereegranskat)abstract In this work we measured, by using a direct approach, the equilibrium solubility of recombinant Aβ40 peptide to be S = 0.36 ± 0.15 μM in aqueous solution of 20 mM sodium phosphate buffer at pH 7.4. Microfluidic diffusional sizing (MDS) and mass spectrometry (MALDI-TOF/TOF) with isotope standard were used to quantify the concentration of soluble Aβ40 species coexisting with formed fibrils. A sample set of Aβ40 at different concentrations was incubated at 37 °C under quiescent conditions and for different incubation times. After incubation, the samples were centrifuged and the concentration of Aβ40 in the supernatant was determined. This allowed us to also determine the metastable zone in supersaturated Aβ40 monomer solutions, as a function of the observation time, as well as the monomer concentration coexisting with formed fibrils. Moreover, the hydrodynamic radius (RH) of the Aβ40 species detected in the supernatant at different time points was also measured, and showed a constant value of ∼1.8 nm, consistent with random coil Aβ40 monomers.
21.	Lima, Tânia, et al. (författare) Understanding the Lipid and Protein Corona Formation on Different Sized Polymeric Nanoparticles 2020 Ingår i: Scientific Reports. - : Springer Science and Business Media LLC. - 2045-2322. ; 10 Tidskriftsartikel (refereegranskat)abstract When in contact with biological fluids, nanoparticles dynamically absorb biomolecules like proteins and lipids onto their surface, forming a “corona”. This biocorona is a dynamic and complex structure that determines how host cells respond to nanoparticles. Despite the common use of mouse models in pre-clinical and toxicological experiments, the impact of corona formed in mouse serum on the biophysical and biological properties of different size NP has not been thoroughly explored. Furthering the knowledge on the corona formed on NP exposed to mouse serum proteins can help in understanding what role it might have in in vivo studies at systemic, tissue, and cellular levels. To investigate biocorona formation, different sized polystyrene NP were exposed to mouse serum. Our data show a size- and time-dependent protein and lipid corona formation. Several proteins were identified and apolipoproteins were by far the most common group on the NPs surfaces. Moreover, we observed that cholesterol and triglycerides effectively bind to NP emphasizing that proteins are not the only biomolecules with high-affinity binding to nanomaterial surfaces. These results highlight that further knowledge on NP interactions with mouse serum is necessary regarding the common use of this model to predict the in vivo efficiency of NP.
22.	Linse, Sara, et al. (författare) Kinetic fingerprints differentiate the mechanisms of action of anti-Aβ antibodies 2020 Ingår i: Nature Structural and Molecular Biology. - : Springer Science and Business Media LLC. - 1545-9993 .- 1545-9985. ; 27:12, s. 1125-1133 Tidskriftsartikel (refereegranskat)abstract The amyloid cascade hypothesis, according to which the self-assembly of amyloid-β peptide (Aβ) is a causative process in Alzheimer’s disease, has driven many therapeutic efforts for the past 20 years. Failures of clinical trials investigating Aβ-targeted therapies have been interpreted as evidence against this hypothesis, irrespective of the characteristics and mechanisms of action of the therapeutic agents, which are highly challenging to assess. Here, we combine kinetic analyses with quantitative binding measurements to address the mechanism of action of four clinical stage anti-Aβ antibodies, aducanumab, gantenerumab, bapineuzumab and solanezumab. We quantify the influence of these antibodies on the aggregation kinetics and on the production of oligomeric aggregates and link these effects to the affinity and stoichiometry of each antibody for monomeric and fibrillar forms of Aβ. Our results reveal that, uniquely among these four antibodies, aducanumab dramatically reduces the flux of Aβ oligomers.
23.	Lundqvist, Martin, et al. (författare) Expression, purification and characterisation of large quantities of recombinant human IAPP for mechanistic studies 2021 Ingår i: Biophysical Chemistry. - : Elsevier BV. - 0301-4622. ; 269 Tidskriftsartikel (refereegranskat)abstract Malfunction and amyloid formation of the Islet Amyloid Polypeptide (IAPP) are factors contributing to Type 2 diabetes. Unravelling the mechanism of IAPP aggregate formation may forward our understanding of this process and its effect on pancreatic β-islet cell. Such mechanistic studies require access to sequence homogeneous and highly pure IAPP. Here we present a new facile protocol for the production of pure recombinant human IAPP at relatively high yield. The protocol uses a His-tagged version of the Npro mutant EDDIE, which drives expression to inclusion bodies, from which the peptide is purified using sonication, refolding and auto-cleavage, removal of EDDIE using Ni-NTA chromatography and reverse-phase HPLC. The purified material is used at multiple concentrations in aggregation kinetics measurements monitored by thioflavin-T fluorescence. Global analysis of the data implies a double nucleation aggregation mechanism including both primary and secondary nucleation.
24.	Mattsson, Karin, et al. (författare) Disaggregation of gold nanoparticles by Daphnia magna 2018 Ingår i: Nanotoxicology. - : Informa UK Limited. - 1743-5390 .- 1743-5404. ; 12:8, s. 885-900 Tidskriftsartikel (refereegranskat)abstract The use of manufactured nanomaterials is rapidly increasing, while our understanding of the consequences of releasing these materials into the environment is still limited and many questions remain, for example, how do nanoparticles affect living organisms in the wild? How do organisms adapt and protect themselves from exposure to foreign materials? How does the environment affect the performance of nanoparticles, including their surface properties? In an effort to address these crucial questions, our main aim has been to probe the effects of aquatic organisms on nanoparticle aggregation. We have, therefore, carried out a systematic study with the purpose to disentangle the effects of the freshwater zooplankter, Daphnia magna, on the surface properties, stability, and aggregation properties of gold (Au) nanoparticles under different aqueous conditions as well as identified the proteins bound to the nanoparticle surface. We show that Au nanoparticles aggregate in pure tap water, but to a lesser extent in water that either contains Daphnia or has been pre-conditioned with Daphnia. Moreover, we show that proteins generated by Daphnia bind to the Au nanoparticles and create a modified surface that renders them less prone to aggregation. We conclude that the surrounding milieu, as well as the surface properties of the original Au particles, are important factors in determining how the nanoparticles are affected by biological metabolism. In a broader context, our results show how nanoparticles released into a natural ecosystem become chemically and physically altered through the dynamic interactions between particles and organisms, either through biological metabolism or through the interactions with biomolecules excreted by organisms into the environment.
25.	Michaels, Thomas C.T., et al. (författare) Dynamics of oligomer populations formed during the aggregation of Alzheimer’s Aβ42 peptide 2020 Ingår i: Nature Chemistry. - : Springer Science and Business Media LLC. - 1755-4330 .- 1755-4349. ; 12:5, s. 445-451 Tidskriftsartikel (refereegranskat)abstract Oligomeric species populated during the aggregation of the Aβ42 peptide have been identified as potent cytotoxins linked to Alzheimer’s disease, but the fundamental molecular pathways that control their dynamics have yet to be elucidated. By developing a general approach that combines theory, experiment and simulation, we reveal, in molecular detail, the mechanisms of Aβ42 oligomer dynamics during amyloid fibril formation. Even though all mature amyloid fibrils must originate as oligomers, we found that most Aβ42 oligomers dissociate into their monomeric precursors without forming new fibrils. Only a minority of oligomers converts into fibrillar structures. Moreover, the heterogeneous ensemble of oligomeric species interconverts on timescales comparable to those of aggregation. Our results identify fundamentally new steps that could be targeted by therapeutic interventions designed to combat protein misfolding diseases. [Figure not available: see fulltext.].
26.	Nilsson, Robert, et al. (författare) Proteomics of Plasma Membranes from Poplar Trees Reveals Tissue Distribution of Transporters, Receptors, and Proteins in Cell Wall Formation 2010 Ingår i: Molecular & Cellular Proteomics. - 1535-9484 .- 1535-9476. ; 9:2, s. 368-387 Tidskriftsartikel (refereegranskat)abstract By exploiting the abundant tissues available from Populus trees, 3-4 m high, we have been able to isolate plasma membranes of high purity from leaves, xylem, and cambium/phloem at a time (4 weeks after bud break) when photosynthesis in the leaves and wood formation in the xylem should have reached a steady state. More than 40% of the 956 proteins identified were found in the plasma membranes of all three tissues and may be classified as "housekeeping" proteins, a typical example being P-type H+-ATPases. Among the 213 proteins predicted to be integral membrane proteins, transporters constitute the largest class (41%) followed by receptors (14%) and proteins involved in cell wall and carbohydrate metabolism (8%) and membrane trafficking (8%). ATP-binding cassette transporters (all members of subfamilies B, C, and G) and receptor-like kinases (four subfamilies) were two of the largest protein families found, and the members of these two families showed pronounced tissue distribution. Leaf plasma membranes were characterized by a very high proportion of transporters, constituting almost half of the integral proteins. Proteins involved in cell wall synthesis (such as cellulose and sucrose synthases) and membrane trafficking were most abundant in xylem plasma membranes in agreement with the role of the xylem in wood formation. Twenty-five integral proteins and 83 soluble proteins were exclusively found in xylem plasma membranes, which identifies new candidates associated with cell wall synthesis and wood formation. Among the proteins uniquely found in xylem plasma membranes were most of the enzymes involved in lignin biosynthesis, which suggests that they may exist as a complex linked to the plasma membrane. Molecular & Cellular Proteomics 9:368-387, 2010.
27.	Pálmadóttir, Tinna, et al. (författare) Morphology-Dependent Interactions between α-Synuclein Monomers and Fibrils 2023 Ingår i: International Journal of Molecular Sciences. - : MDPI AG. - 1661-6596 .- 1422-0067. ; 24:6 Tidskriftsartikel (refereegranskat)abstract Amyloid fibrils may adopt different morphologies depending on the solution conditions and the protein sequence. Here, we show that two chemically identical but morphologically distinct α-synuclein fibrils can form under identical conditions. This was observed by nuclear magnetic resonance (NMR), circular dichroism (CD), and fluorescence spectroscopy, as well as by cryo-transmission electron microscopy (cryo-TEM). The results show different surface properties of the two morphologies, A and B. NMR measurements show that monomers interact differently with the different fibril surfaces. Only a small part of the N-terminus of the monomer interacts with the fibril surface of morphology A, compared to a larger part of the monomer for morphology B. Differences in ThT binding seen by fluorescence titrations, and mesoscopic structures seen by cryo-TEM, support the conclusion of the two morphologies having different surface properties. Fibrils of morphology B were found to have lower solubility than A. This indicates that fibrils of morphology B are thermodynamically more stable, implying a chemical potential of fibrils of morphology B that is lower than that of morphology A. Consequently, at prolonged incubation time, fibrils of morphology B remained B, while an initially monomorphic sample of morphology A gradually transformed to B.
28.	Rodriguez Camargo, Diana C., et al. (författare) Proliferation of Tau 304-380 Fragment Aggregates through Autocatalytic Secondary Nucleation 2021 Ingår i: ACS Chemical Neuroscience. - : American Chemical Society (ACS). - 1948-7193. ; 12:23, s. 4406-4415 Tidskriftsartikel (refereegranskat)abstract The self-assembly of the protein tau into neurofibrillary tangles is one of the hallmarks of Alzheimer's disease and related tauopathies. Still, the molecular mechanism of tau aggregation is largely unknown. This problem may be addressed by systematically obtaining reproducible in vitro kinetics measurements under quiescent conditions in the absence of triggering substances. Here, we implement this strategy by developing protocols for obtaining an ultrapure tau fragment (residues 304-380 of tau441) and for performing spontaneous aggregation assays with reproducible kinetics under quiescent conditions. We are thus able to identify the mechanism of fibril formation of the tau 304-380 fragment at physiological pH using fluorescence spectroscopy and mass spectrometry. We find that primary nucleation is slow, and that secondary processes dominate the aggregation process once the initial aggregates are formed. Moreover, our results further show that secondary nucleation of monomers on fibril surfaces dominates over fragmentation of fibrils. Using separate isotopes in monomers and fibrils, through mass spectroscopy measurements, we verify the isotope composition of the intermediate oligomeric species, which reveals that these small aggregates are generated from monomer through secondary nucleation. Our results provide a framework for understanding the processes leading to tau aggregation in disease and for selecting possible tau forms as targets in the development of therapeutic interventions in Alzheimer's disease.
29.	Rodriguez Camargo, Diana C., et al. (författare) Surface-Catalyzed Secondary Nucleation Dominates the Generation of Toxic IAPP Aggregates 2021 Ingår i: Frontiers in Molecular Biosciences. - : Frontiers Media SA. - 2296-889X. ; 8 Tidskriftsartikel (refereegranskat)abstract The aggregation of the human islet amyloid polypeptide (IAPP) is associated with diabetes type II. A quantitative understanding of this connection at the molecular level requires that the aggregation mechanism of IAPP is resolved in terms of the underlying microscopic steps. Here we have systematically studied recombinant IAPP, with amidated C-terminus in oxidised form with a disulphide bond between residues 3 and 7, using thioflavin T fluorescence to monitor the formation of amyloid fibrils as a function of time and IAPP concentration. We used global kinetic analyses to connect the macroscopic measurements of aggregation to the microscopic mechanisms, and show that the generation of new aggregates is dominated by the secondary nucleation of monomers on the fibril surface. We then exposed insulinoma cells to aliquots extracted from different time points of the aggregation process, finding the highest toxicity at the midpoint of the reaction, when the secondary nucleation rate reaches its maximum. These results identify IAPP oligomers as the most cytotoxic species generated during IAPP aggregation, and suggest that compounds that target secondary nucleation of IAPP could be most effective as therapeutic candidates for diabetes type II.
30.	Söderberg, Christopher A.G., et al. (författare) Structural modelling of the DNAJB6 oligomeric chaperone shows a peptide-binding cleft lined with conserved S/T-residues at the dimer interface 2018 Ingår i: Scientific Reports. - : NATURE PUBLISHING GROUP. - 2045-2322. ; 8 Tidskriftsartikel (refereegranskat)abstract = The remarkably efficient suppression of amyloid fibril formation by the DNAJB6 chaperone is dependent on a set of conserved S/T-residues and an oligomeric structure, features unusual among DNAJ chaperones. We explored the structure of DNAJB6 using a combination of structural methods. Lysine-specific crosslinking mass spectrometry provided distance constraints to select a homology model of the DNAJB6 monomer, which was subsequently used in crosslink-assisted docking to generate a dimer model. A peptide-binding cleft lined with S/T-residues is formed at the monomer-monomer interface. Mixed isotope crosslinking showed that the oligomers are dynamic entities that exchange subunits. The purified protein is well folded, soluble and composed of oligomers with a varying number of subunits according to small-angle X-ray scattering (SAXS). Elongated particles (160 x 120 angstrom) were detected by electron microscopy and single particle reconstruction resulted in a density map of 20 angstrom resolution into which the DNAJB6 dimers fit. The structure of the oligomer and the S/T-rich region is of great importance for the understanding of the function of DNAJB6 and how it can bind aggregation-prone peptides and prevent amyloid diseases.
31.	Tran, Huy Cuong, et al. (författare) An mTRAN-mRNA interaction mediates mitochondrial translation initiation in plants 2023 Ingår i: Science. - 1095-9203. ; 381:6661 Tidskriftsartikel (refereegranskat)abstract Plant mitochondria represent the largest group of respiring organelles on the planet. Plant mitochondrial messenger RNAs (mRNAs) lack Shine-Dalgarno-like ribosome-binding sites, so it is unknown how plant mitoribosomes recognize mRNA. We show that “mitochondrial translation factors” mTRAN1 and mTRAN2 are land plant–specific proteins, required for normal mitochondrial respiration chain biogenesis. Our studies suggest that mTRANs are noncanonical pentatricopeptide repeat (PPR)–like RNA binding proteins of the mitoribosomal “small” subunit. We identified conserved Adenosine (A)/Uridine (U)-rich motifs in the 5′ regions of plant mitochondrial mRNAs. mTRAN1 binds this motif, suggesting that it is a mitoribosome homing factor to identify mRNAs. We demonstrate that mTRANs are likely required for translation of all plant mitochondrial mRNAs. Plant mitochondrial translation initiation thus appears to use a protein-mRNA interaction that is divergent from bacteria or mammalian mitochondria.
32.	Wagner-Egea, Paula, et al. (författare) Marine PET Hydrolase (PET2) : Assessment of Terephthalate- and Indole-Based Polyester Depolymerization 2023 Ingår i: Catalysts. - 2073-4344. ; 13:9 Tidskriftsartikel (refereegranskat)abstract Enzymatic polyethylene terephthalate (PET) recycling processes are gaining interest for their low environmental impact, use of mild conditions, and specificity. Furthermore, PET hydrolase enzymes are continuously being discovered and engineered. In this work, we studied a PET hydrolase (PET2), initially characterized as an alkaline thermostable lipase. PET2 was produced in a fusion form with a 6-histidine tag in the N-terminal. The PET2 activity on aromatic terephthalate and new indole-based polyesters was evaluated using polymers in powder form. Compared with IsPETase, an enzyme derived from Ideonella sakaiensis, PET2 showed a lower PET depolymerization yield. However, interestingly, PET2 produced significantly higher polybutylene terephthalate (PBT) and polyhexylene terephthalate (PHT) depolymerization yields. A clear preference was found for aromatic indole-derived polyesters over non-aromatic ones. No activity was detected on Akestra™, an amorphous copolyester with spiroacetal structures. Docking studies suggest that a narrower and more hydrophobic active site reduces its activity on PET but favors its interaction with PBT and PHT. Understanding the enzyme preferences of polymers will contribute to their effective use to depolymerize different types of polyesters.

Skapa referenser, mejla, bekava och länka

Länka till träfflistan

Träfflista för sökning "WFRF:(Bernfur Katja) "

Avgränsa träffmängd

År