Cloning and expression of a transferrin-binding protein from Actinobacillus pleuropneumoniae
| Autor: | Sandy Klashinsky, Gerald-F. Gerlach, Philip Willson, Andrew A. Potter, C Anderson |
|---|---|
| Rok vydání: | 1992 |
| Předmět: |
Swine
Immunology Heme Molecular cloning medicine.disease_cause Microbiology law.invention chemistry.chemical_compound law Iron-Binding Proteins Immunoscreening Escherichia coli medicine Animals Cloning Molecular Actinobacillus pleuropneumoniae chemistry.chemical_classification biology Binding protein Transferrin Transferrin-Binding Proteins biology.organism_classification Molecular biology Infectious Diseases chemistry Genes Bacterial Recombinant DNA Parasitology Transformation Bacterial Carrier Proteins Research Article Hemin |
| Zdroj: | Infection and Immunity. 60:892-898 |
| ISSN: | 1098-5522 0019-9567 |
| DOI: | 10.1128/iai.60.3.892-898.1992 |
| Popis: | An expression library was constructed from Actinobacillus pleuropneumoniae serotype 7. Escherichia coli transformants expressing recombinant proteins were identified by immunoscreening with porcine convalescent serum. One transformant expressing a 60-kDa protein (60K protein) in aggregated form was identified. Serum raised against the recombinant protein recognized a polypeptide with an indistinguishable electrophoretic mobility in the A. pleuropneumoniae wild type after iron-restricted growth only. The recombinant protein bound transferrin after blotting onto nitrocellulose. Using a competitive enzyme-linked immunosorbent assay (ELISA), the specificity of this binding for the amino-terminal half of iron-saturated porcine transferrin was established. Also, the 60K wild-type protein bound hemin as assessed by hemin-agarose chromatography. Hemin could inhibit transferrin binding of the recombinant protein in the competitive ELISA, whereas hemoglobin and synthetic iron chelators failed to do so. Southern blot analysis of several other A. pleuropneumoniae strains indicated that highly homologous sequence is present in eight of eight isolates of serotype 7 and in some isolates of serotypes 2, 3, and 4. |
| Databáze: | OpenAIRE |
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