Search results for "Protein multimerization"
showing 10 items of 99 documents
Wild-type Cu/Zn superoxide dismutase (SOD1) does not facilitate, but impedes the formation of protein aggregates of amyotrophic lateral sclerosis cau…
2009
Aggregation of Cu/Zn superoxide dismutase (SOD1) is a hallmark of a subset of familial amyotrophic lateral sclerosis (ALS) cases. The expression of wild-type SOD1 [SOD(hWT)] surprisingly exacerbates the phenotype of mutant SOD1 in vivo. Here we studied whether SOD1(hWT) may affect mutant SOD1 aggregation by employing fluorescence microscopy techniques combined with lifetime-based Förster resonance energy transfer (FRET). Only a very minor fraction of SOD1(hWT) was observed in aggregates induced by mutant SOD1(G37R), SOD1(G85R) or SOD1(G93C). Quite in contrast, co-expression of SOD(hWT) reduced the amount of mutant SOD1 in the aggregate fraction. Furthermore, we did not detect endogenous mou…
Polar/Ionizable Residues in Transmembrane Segments: Effects on Helix-Helix Packing
2012
The vast majority of membrane proteins are anchored to biological membranes through hydrophobic alpha-helices. Sequence analysis of high-resolution membrane protein structures show that ionizable amino acid residues are present in transmembrane (TM) helices, often with a functional and/or structural role. Here, using as scaffold the hydrophobic TM domain of the model membrane protein glycophorin A (GpA), we address the consequences of replacing specific residues by ionizable amino acids on TM helix insertion and packing, both in detergent micelles and in biological membranes. Our findings demonstrate that ionizable residues are stably inserted in hydrophobic environments, and tolerated in t…
Thioflavin T templates amyloid β(1–40) conformation and aggregation pathway
2015
Aβ(1-40) peptide supramolecular assembly and fibril formation processes are widely recognized to have direct implications in the progression of Alzheimer's disease. The molecular basis of this biological process is still unknown and there is a strong need of developing effective strategies to control the occurring events. To this purpose the exploitation of small molecules interacting with Aβ aggregation represents one of the possible routes. Moreover, the use specific labeling has represented so far one of the most common and effective methods to investigate such a process. This possibility in turn rests on the reliability of the probe/labels involved. Here we present evidences of the effe…
Mutational analyses of YqjA, a Tvp38/DedA protein of E. coli
2015
AbstractMembrane proteins of the DedA/Tvp38 protein family are involved in membrane integrity and virulence of pathogenic organisms. However, the structure and exact function of any member of this large protein family are still unclear. In the present study we analyzed the functional and structural properties of a DedA homolog. Purified YqjA variants from Escherichia coli are detectable in different oligomeric states and specific homo-interaction of YqjA monomers in the membrane were confirmed by formation of a disulfide bond in the C-terminal transmembrane helix. Moreover, alanine scanning mutagenesis exhibited different interaction sites crucial for YqjA activity vs. dimer formation.
Heat shock protein 27 is involved in SUMO-2/3 modification of heat shock factor 1 and thereby modulates the transcription factor activity
2009
Heat shock protein 27 (HSP27) accumulates in stressed cells and helps them to survive adverse conditions. We have already shown that HSP27 has a function in the ubiquitination process that is modulated by its oligomerization/phosphorylation status. Here, we show that HSP27 is also involved in protein sumoylation, a ubiquitination-related process. HSP27 increases the number of cell proteins modified by small ubiquitin-like modifier (SUMO)-2/3 but this effect shows some selectivity as it neither affects all proteins nor concerns SUMO-1. Moreover, no such alteration in SUMO-2/3 conjugation is achievable by another HSP, such as HSP70. Heat shock factor 1 (HSF1), a transcription factor responsib…
Human apolipoprotein A-I natural variants: molecular mechanisms underlying amyloidogenic propensity
2012
Human apolipoprotein A-I (apoA-I)-derived amyloidosis can present with either wild-type (Wt) protein deposits in atherosclerotic plaques or as a hereditary form in which apoA-I variants deposit causing multiple organ failure. More than 15 single amino acid replacement amyloidogenic apoA-I variants have been described, but the molecular mechanisms involved in amyloid-associated pathology remain largely unknown. Here, we have investigated by fluorescence and biochemical approaches the stabilities and propensities to aggregate of two disease-associated apoA-I variants, apoA-IGly26Arg, associated with polyneuropathy and kidney dysfunction, and apoA-ILys107-0, implicated in amyloidosis in severe…
Cryopreservation of MHC Multimers: Recommendations for Quality Assurance in Detection of Antigen Specific T Cells
2015
Fluorescence-labeled peptide-MHC class I multimers serve as ideal tools for the detection of antigen-specific T cells by flow cytometry, enabling functional and phenotypical characterization of specific T cells at the single cell level. While this technique offers a number of unique advantages, MHC multimer reagents can be difficult to handle in terms of stability and quality assurance. The stability of a given fluorescence-labeled MHC multimer complex depends on both the stability of the peptide-MHC complex itself and the stability of the fluorochrome. Consequently, stability is difficult to predict and long-term storage is generally not recommended. We investigated here the possibility of…
Lipid and phase specificity of α-toxin from S. aureus
2013
AbstractThe pore forming toxin Hla (α-toxin) from Staphylococcus aureus is an important pathogenic factor of the bacterium S. aureus and also a model system for the process of membrane-induced protein oligomerisation and pore formation. It has been shown that binding to lipid membranes at neutral or basic pH requires the presence of a phosphocholine-headgroup. Thus, sphingomyelin and phosphatidylcholine may serve as interaction partners in cellular membranes. Based on earlier studies it has been suggested that rafts of sphingomyelin are particularly efficient in toxin binding. In this study we compared the oligomerisation of Hla on liposomes of various lipid compositions in order to identif…
Thermal Aggregation of Bovine Serum Albumin in Trehalose and Sucrose Aqueous Solutions
2012
We report results of static and dynamic light scattering measurements performed on bovine serum albumin (BSA) in saccharide (trehalose and sucrose) solutions. Our aim is to study the effects of the two disaccharides on the first steps of thermal aggregation of BSA in aqueous solutions at two protein concentrations (1 and 30 mg/mL) at increasing sugar/water ratio. Results show that sugars modify early stages of aggregation mainly by perturbing the thermodynamic behavior of the solvent (i.e., general solvent effects) without involving direct, specific sugar-protein interactions. This agrees with current hypotheses on sugar action in protein solutions. (1-3) The linear correlation detected bet…
Dimerization of visinin-like protein 1 is regulated by oxidative stress and calcium and is a pathological hallmark of amyotrophic lateral sclerosis
2014
AbstractRedox control of proteins that form disulfide bonds upon oxidative challenge is an emerging topic in the physiological and pathophysiological regulation of protein function. We have investigated the role of the neuronal calcium sensor protein visinin-like protein 1 (VILIP-1) as a novel redox sensor in a cellular system. We have found oxidative stress to trigger dimerization of VILIP-1 within a cellular environment and identified thioredoxin reductase as responsible for facilitating the remonomerization of the dimeric protein. Dimerization is modulated by calcium and not dependent on the myristoylation of VILIP-1. Furthermore, we show by site-directed mutagenesis that dimerization is…