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A model of the rabies virus glycoprotein active site
No full textThe glycoprotein from the neurotropic rabies virus shows a significant homology with the alpha neurotoxin that binds to the nicotinic acetylcholine receptor. The crystal structure of the alpha neurotoxins suggests that the Arg 37 guanidinium group and the Asp 31 side-chain carboxylate of the erabutoxin have stereochemical features resembling those of acetylcholine. Conformational studies on the Asn194-Ser195-Arg196-Gly197 tetrapeptide, an essential part of the binding site of the rabies virus glycoprotein, indicate that the side chains of Asn and Arg could also mimic the acetylcholine structure. This observation is consistent with the recently proposed mechanism of the viral infection
High performance liquid chromatography immunoaffinity purification of antibodies and antibody fragments
No full textA rapid antibody purification method is described which combines high performance liquid chromatography (HPLC) resolution with affinity chromatography specificity. The antigen used as immobilized ligand is bound to the HPLC column matrix by reacting the amino groups of the protein with the active epoxy groups of the latter. Once the reaction has finished, the unreacted groups of the column are saturated with an appropriate scavenger. Unrelated proteins are then washed out and the specific antibodies recovered by lowering the pH of the elution buffer
Micro-determination of amino acid composition of proteins electroblotted onto polyvinylidene difluoride membranes
No full textA method for determination of amino acid composition of proteins separated by SDS-polyacrylamide gel electrophoresis and transferred to polyvinylidene difluoride (PVDF) membranes is described. A single blotted band containing 50 to 200 pmoles of protein was cut out and submitted to acid hydrolysis with HCl followed by derivatization with phenylisothiocyanate. The amino acid derivatives were separated by reverse phase high-performance liquid chromatography. Bovine serum albumin, lysozyme, myoglobin, ovalbumin, soybean trypsin inhibitor and carbonic anhydrase were analyzed; the results revealed a good correspondence with reported values. This can be considered an analytical method to determine the amino acid composition of samples from microquantities of protein mixtures, particularly in those cases in which SDS-polyacrylamide gel electrophoresis is the most suitable separation system
Immunoproteomics of myasthenia
No full textAutoantibodies to nAChR are found in 85% of patients with Myasthenia Gravis (seropositive) while in 15% of them the standard immunoprecipitation test for anti-AChR is negative (seronegative). Some of these seronegative patients have antibodies against muscle specific Kinase ( MuSK). The aim of this study is to try to define auto-antibodies against protein different from AChR and MuSK in seronegative patients. We used the cell line TE671, that expresses human AChR. Through our proteomic approach we identified some proteins that could represent potential antigens different from AChR and MuSK
A monoclonal antibody to a synthetic fragment of rabies virus glycoprotein binds ligands of the nicotinic cholinergic receptor
No full textRabies virus glycoprotein and snake venom curaremimetic neurotoxins share a region of high homology (30–45 for neurotoxins and 190–203 for the glycoprotein) in the regions that are believed to be responsible for binding the nicotinic acetylcholine receptor. Monoclonal antibodies raised to the 190–203 synthetic fragment of rabies virus glycoprotein were immobilized on a high performance affinity chromatography column and were able to bind neurotoxins. Toxins were displaced from the affinity column by elution at acidic pH and by affinity competition with acetylcholine at neutral pH. Furthermore, the affinity column proved to be useful for the purification of cholinergic ligands. Overall, these results indicate that the paratope of our monoclonal antibodies could behave as an ‘internal image’ of the nicotinic cholinergic receptor acetylcholine binding site
Purification of acidic synthetic peptides by high performance liquid chromatography using ammonium acetate buffer
No full textThe purification of acidic synthetic peptides was achieved by using a linear gradient of ammonium acetate and methanol with reverse phase liquid chromatography
Going Beyond Counting First Authors in Author Co-citation Analysis
Full text linkThe present study examines one of the fundamental aspects of author co-citation analysis (ACA) - the way co-citation
counts are defined. Co-citation counting provides the data on which all subsequent statistical analyses and mappings
are based, and we compare ACA results based on two different types of co-citation counting - the traditional type that
only counts the first one among a cited work's authors on the one hand and a non-traditional type that takes into
account the first 5 authors of a cited work on the other hand. Results indicate that the picture produced through this non-traditional author co-citation counting contains more coherent author groups and is therefore considerably clearer. However, this picture represents fewer specialties in the research field being studied than that produced through the traditional first-author co-citation counting when the same number of top-ranked authors is selected and analyzed. Reasons for these effects are discussed
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