Abstract
The present study characterized the biological function of the asparaginyl peptidase legumain-1 (LEG-1) of the bovine lungworm Dictyocaulus viviparus and its suitability as a recombinant vaccine against dictyocaulosis. Quantitative real-time PCR and immunoblot analysis revealed LEG-1 to be almost exclusively transcribed and expressed in parasitic lungworm stages. Immunohistochemistry localized the enzyme in the parasite's gut, which was confirmed by immunoblots detecting LEG-1 in the gut as well as male testes. LEG-1 was recombinantly (rLEG-1) expressed in the yeast Pichia pastoris and subsequently analysed in activity assays for its enzyme functions and substrate specificity. For sufficient functionality, rLEG-1 needed trans-activation through D. viviparus cathepsin L-2, indicating a novel mechanism of legumain activation. After trans-activation, rLEG-1 worked best at pH 5.5 and 35-39 degrees C and cleaved a legumain-specific artificial substrate as well as the natural substrates bovine collagen types I and II. In a clinical vaccination trial, rLEG-1 did not protect against challenge infection. Results of in vitro characterization, transcription pattern and localization enhance the presumption that LEG-1 participates in digestion processes of D. viviparus. Since rLEG-1 needs trans-activation through a cathepsin, it is probably involved in an enzyme cascade and therefore remains interesting as a candidate in a multi-component vaccine.
| Dokumententyp: | Zeitschriftenartikel |
|---|---|
| Fakultät: | Biologie > Department Biologie II |
| Themengebiete: | 500 Naturwissenschaften und Mathematik > 570 Biowissenschaften; Biologie |
| ISSN: | 0031-1820 |
| Sprache: | Englisch |
| Dokumenten ID: | 67710 |
| Datum der Veröffentlichung auf Open Access LMU: | 19. Jul. 2019, 12:23 |
| Letzte Änderungen: | 04. Nov. 2020, 13:49 |
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