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| Alternative Name : |
Outer Membrane Protein-A, OmpA. |
| Amount : |
200 µg |
Source : Escherichia Coli. The OmpA protein is one of the main outer-membrane proteins of a large array of Gram-negative bacteria such as A.salmonicida, Shigella dysenteriae and E.coli.OmpA's major physiological functions include maintenance of the structural integrity and morphology of the cells and porin activity, as well as a role in conjugation and bacteriophage binding.Achromogenic atypical Aeromonas salmonicida is the causative agent of goldfish ulcer disease.Virulence of this bacterium is associated with the production of a paracrystalline outer membrane A-layer protein.The species specific structural gene for the monomeric form of A-protein was cloned into a pET-3d plasmid in order to express and produce a recombinant form of the protein in E.coli BL21(DE3). The induced protein was isolated from inclusion bodies by a simple solubilization-renaturation procedure and purified by ion exchange chromatography on Q-Sepharose to over 95% pure monomeric protein.Recombinant A-protein was compared by biochemical, immunological and molecular methods with the A-protein isolated from atypical A.salmonicida bacterial cells by the glycine and the membrane extraction methods.
It is recommended to reconstitute the lyophilized OmpA in sterile 0.4% NaHCO3, not less than 100µg/ml, which can then be further diluted to other aqueous solutions. The interaction of bacterial and recombinant A-layer protein with murine macrophages was directed at determining the effect of A-protein on intracellular events that occur in primed macrophages. This was accomplished by measuring the cytotoxic product produced by peritoneal macrophages when exposed to A-protein coated latex beads. Thioglycolate elicited macrophages exhibited a low level of activation (18% cytotoxicity) that was significantly increased (48% cytotoxicity) in the presence of latex beads. Coating of the latex beads with each of the three A-protein products resulted in an increase of cytoxicity (mean +/- SEM) from 48% to 91%.
