363 research outputs found
Does McLuhan’s Idea Stand Up for Millennials? Testing Whether the Medium is the Message in Political Organization Public Relationships
Examining McLuhan’s (1964) classic adage that the “medium is the message,” this experimental
design presented participants with the three versions of the same message content. Manipulating
medium as an online press release, blog post, or online video, the research here found that
medium does play a role in shaping the receiver’s perceived relationship with an organization.
Conversely, medium does not impact one’s assessment of credibility. Set in a digital political
public relations context, this study attempts to connect public relations’ most heuristic theory in
the examination in the state of ever-evolving digital media
Folding of the Protein Domain hbSBD
The folding of the alpha-helice domain hbSBD of the mammalian mitochondrial
branched-chain alpha-ketoacid dehydrogenase (BCKD) complex is studied by the
circular dichroism technique in absence of urea. Thermal denaturation is used
to evaluate various thermodynamic parameters defining the equilibrium
unfolding, which is well described by the two-state model with the folding
temperature T_f = 317.8 K and the enthalpy change Delta H_g = 19.67 kcal/mol.
The folding is also studied numerically using the off-lattice coarse-grained Go
model and the Langevin dynamics. The obtained results, including the population
of the native basin, the free energy landscape as a function of the number of
native contacts and the folding kinetics, also suggest that the hbSBD domain is
a two-state folder. These results are consistent with the biological function
of hbSBD in BCKD.Comment: 25 pages, 7 figures, 1 table, published in Biophysical Journa
Mathematics, Thermodynamics, and Modeling to Address Ten Common Misconceptions about Protein Structure, Folding, and Stability
To fully understand the roles proteins play in cellular processes, students need to grasp complex ideas about protein structure, folding, and stability. Our current understanding of these topics is based on mathematical models and experimental data. However, protein structure, folding, and stability are often introduced as descriptive, qualitative phenomena in undergraduate classes. In the process of learning about these topics, students often form incorrect ideas. For example, by learning about protein folding in the context of protein synthesis, students may come to an incorrect conclusion that once synthesized on the ribosome, a protein spends its entire cellular life time in its fully folded native confirmation. This is clearly not true; proteins are dynamic structures that undergo both local fluctuations and global unfolding events. To prevent and address such misconceptions, basic concepts of protein science can be introduced in the context of simple mathematical models and hands-on explorations of publicly available data sets. Ten common misconceptions about proteins are presented, along with suggestions for using equations, models, sequence, structure, and thermodynamic data to help students gain a deeper understanding of basic concepts relating to protein structure, folding, and stability
Prots: A fragment based protein thermo‐stability potential
Designing proteins with enhanced thermo‐stability has been a main focus of protein engineering because of its theoretical and practical significance. Despite extensive studies in the past years, a general strategy for stabilizing proteins still remains elusive. Thus effective and robust computational algorithms for designing thermo‐stable proteins are in critical demand. Here we report PROTS, a sequential and structural four‐residue fragment based protein thermo‐stability potential. PROTS is derived from a nonredundant representative collection of thousands of thermophilic and mesophilic protein structures and a large set of point mutations with experimentally determined changes of melting temperatures. To the best of our knowledge, PROTS is the first protein stability predictor based on integrated analysis and mining of these two types of data. Besides conventional cross validation and blind testing, we introduce hypothetical reverse mutations as a means of testing the robustness of protein thermo‐stability predictors. In all tests, PROTS demonstrates the ability to reliably predict mutation induced thermo‐stability changes as well as classify thermophilic and mesophilic proteins. In addition, this white‐box predictor allows easy interpretation of the factors that influence mutation induced protein stability changes at the residue level. Proteins 2012; © 2011 Wiley Periodicals, Inc.Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/89526/1/23163_ftp.pd
Non-native hydrophobic interactions detected in unfolded apoflavodoxin by paramagnetic relaxation enhancement
Transient structures in unfolded proteins are important in elucidating the molecular details of initiation of protein folding. Recently, native and non-native secondary structure have been discovered in unfolded A. vinelandii flavodoxin. These structured elements transiently interact and subsequently form the ordered core of an off-pathway folding intermediate, which is extensively formed during folding of this α–β parallel protein. Here, site-directed spin-labelling and paramagnetic relaxation enhancement are used to investigate long-range interactions in unfolded apoflavodoxin. For this purpose, glutamine-48, which resides in a non-native α-helix of unfolded apoflavodoxin, is replaced by cysteine. This replacement enables covalent attachment of nitroxide spin-labels MTSL and CMTSL. Substitution of Gln-48 by Cys-48 destabilises native apoflavodoxin and reduces flexibility of the ordered regions in unfolded apoflavodoxin in 3.4 M GuHCl, because of increased hydrophobic interactions in the unfolded protein. Here, we report that in the study of the conformational and dynamic properties of unfolded proteins interpretation of spin-label data can be complicated. The covalently attached spin-label to Cys-48 (or Cys-69 of wild-type apoflavodoxin) perturbs the unfolded protein, because hydrophobic interactions occur between the label and hydrophobic patches of unfolded apoflavodoxin. Concomitant hydrophobic free energy changes of the unfolded protein (and possibly of the off-pathway intermediate) reduce the stability of native spin-labelled protein against unfolding. In addition, attachment of MTSL or CMTSL to Cys-48 induces the presence of distinct states in unfolded apoflavodoxin. Despite these difficulties, the spin-label data obtained here show that non-native contacts exist between transiently ordered structured elements in unfolded apoflavodoxin
Physicochemical and biological characterization of 1E10 Anti-Idiotype vaccine
<p>Abstract</p> <p>Background</p> <p>1E10 monoclonal antibody is a murine anti-idiotypic antibody that mimics N-glycolyl-GM3 gangliosides. This antibody has been tested as an anti-idiotypic cancer vaccine, adjuvated in Al(OH)<sub>3</sub>, in several clinical trials for melanoma, breast, and lung cancer. During early clinical development this mAb was obtained <it>in vivo </it>from mice ascites fluid. Currently, the production process of 1E10 is being transferred from the <it>in vivo </it>to a bioreactor-based method.</p> <p>Results</p> <p>Here, we present a comprehensive molecular and immunological characterization of 1E10 produced by the two different production processes in order to determine the impact of the manufacturing process in vaccine performance. We observed differences in glycosylation pattern, charge heterogeneity and structural stability between <it>in vivo</it>-produced 1E10 and bioreactor-obtained 1E10. Interestingly, these modifications had no significant impact on the immune responses elicited in two different animal models.</p> <p>Conclusions</p> <p>Changes in 1E10 primary structure like glycosylation; asparagine deamidation and oxidation affected 1E10 structural stability but did not affect the immune response elicited in mice and chickens when compared to 1E10 produced in mice.</p
Structural and functional stabilization of protein entities: state-of-the-art
Within the context of biomedicine and pharmaceutical sciences, the issue of (therapeutic) protein stabilization assumes particular relevance. Stabilization of protein and protein-like molecules translates into preservation of both structure and functionality during storage and/or targeting, and such stabilization is mostly attained through establishment of a thermodynamic equilibrium with the (micro)environment. The basic thermodynamic principles that govern protein structural transitions and the interactions of the protein molecule with its (micro)environment are, therefore, tackled in a systematic fashion. Highlights are given to the major classes of (bio)therapeutic molecules, viz. enzymes, recombinant proteins, (macro)peptides, (monoclonal) antibodies and bacteriophages. Modification of the microenvironment of the biomolecule via multipoint covalent attachment onto a solid surface followed by hydrophilic polymer co-immobilization, or physical containment within nanocarriers, are some of the (latest) strategies discussed aiming at full structural and functional stabilization of said biomolecules.Financial support to Victor M. Balcao, via an Invited Research Scientist fellowship (FAPESP Ref. No. 2011/51077-8), and project funding (FAPESP Ref. No. 2013/03181-6, Project PneumoPhageKill) by Fundacao de Amparo a Pesquisa do Estado de Sao Paulo (FAPESP, Sao Paulo, Brazil), is hereby gratefully acknowledged. The authors are also grateful to Claudio M. Barroso (BSc.), Graphic Designer at University of Sorocaba (UNISO), for computer-designing the schemes/drawings integrating this review paper
Common Features at the Start of the Neurodegeneration Cascade
A single-molecule study reveals that neurotoxic proteins share common structural features that may trigger neurodegeneration, thus identifying new targets for therapy and diagnosis
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Targeting Lipid Metabolism in Myeloid Cells to Improve Ischemic Stroke Recovery
Globally, more than 67 million people are living with the effects of ischemic stroke. Importantly, many stroke survivors develop a chronic inflammatory response that may contribute to cognitive impairment, a common and debilitating sequela of stroke that is insufficiently studied and currently untreatable. Therefore, the goal of my research has been to develop a novel therapeutic to attenuate chronic inflammation and prevent post-stroke cognitive decline in a preclinical murine model of experimental ischemic stroke.
We hypothesize that, following ischemic stroke, lipids derived from myelin debris and other apoptotic or necrotic cell membranes overwhelm the processing capability of infiltrating monocytes and resident microglia in the brain, leading to the formation of lipid-laden foamy macrophages, generation of cholesterol crystals, secretion of pro-inflammatory cytokines, and production of degradative enzymes. Further, we hypothesize that the resultant chronic inflammatory response, coupled with concurrent cell death, causes secondary neurodegeneration and impairs cognitive and locomotor function. To address these hypotheses, the aims of the studies presented herein were three-fold: (1) to characterize the lipidome of chronic stroke infarcts and determine whether the pathogenic lipids are derived from myelin; (2) to determine whether the ablation of lipid metabolic pathways (CD36, NLRP3, C3) impacts stroke recovery; and (3) to determine whether lipid complexation and macrophage reprogramming within infarcts, via the repeated administration of 2-hydroxypropyl-β-cyclodextrin (HPβCD), attenuates chronic inflammation and improves recovery after experimental stroke.
We first demonstrate that lipid metabolism is disrupted in chronic stroke infarcts, which causes the accumulation of uncleared lipid debris and correlates with chronic inflammation. Specifically, we illustrate that, coincident to the infiltration of peripheral immune cells, stroke infarcts amass lipids derived from myelin membranes, including sulfatides, sphingomyelins, fatty acids, and cholesterol esters. To our knowledge, these substantial alterations in lipid homeostasis have not been previously recognized or investigated in the context of ischemic stroke.
We next investigate the roles of lipid metabolic pathways in ischemic stroke recovery. Specifically, we demonstrate that genetic ablation of Cd36 reduces the accumulation of B lymphocytes and improves locomotor function in the weeks after stroke. We also illustrate that the genetic ablation of Nlrp3 or C3 has negligible effects on chronic inflammation at 7 weeks after stroke; although, mice deficient in Nlrp3 demonstrate marginal improvements in tests of spontaneous locomotor function. Together, these results indicate that Cd36 and, to a lesser extent, Nlrp3 have integral roles in chronic inflammation and locomotor function after ischemic stroke.
We then provide a proof of principle that solubilizing and entrapping lipophilic substances using 2-hydroxypropyl-β-cyclodextrin (HPβCD) could be an effective strategy for treating chronic inflammation after ischemic stroke or other central nervous system (CNS) injuries. HPβCD is a U.S. Food and Drug Administration (FDA)-approved cyclic oligosaccharide that promotes liver X receptor (LXR)-mediated transcriptional reprogramming in macrophages and incites anti-inflammatory mechanisms. We illustrate that the repeated administration of HPβCD curtails the chronic inflammatory response to ischemic stroke by reducing lipid accumulation within stroke infarcts. In these preclinical trials, we subcutaneously injected young adult and aged male mice with vehicle or HPβCD three times per week, with treatment beginning one week after ischemic stroke. We demonstrate that chronic stroke infarct and peri-infarct regions in HPβCD-treated mice were characterized by an upregulation of genes involved in lipid metabolism and a downregulation of genes involved in innate and adaptive immunity, reactive astrogliosis, and chemotaxis. Correspondingly, HPβCD reduced the accumulation of lipid droplets, T lymphocytes, B lymphocytes, and plasma cells in stroke infarcts. Repeated administration of HPβCD also preserved NeuN immunoreactivity in the striatum and thalamus and c-Fos immunoreactivity in hippocampal regions. Additionally, HPβCD improved recovery through the protection of hippocampal-dependent spatial working memory and reduction of impulsivity. These results indicate that systemic HPβCD treatment attenuates chronic inflammation and secondary neurodegeneration and prevents post-stroke cognitive decline in a murine model of ischemic stroke. We propose that repurposing HPβCD for the prevention of post-stroke dementia could improve recovery and increase long-term quality of life in stroke sufferers
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