Details zur Publikation

Referenztyp Zeitschriften
DOI / URL Link
Titel (primär) Detection, quantification and identification of fungal extracellular laccases using polyclonal antibody and mass spectrometry
Autor Kellner, H.; Jehmlich, N.; Benndorf, D.; Hoffmann, R.; Rühl, M.; Hoegger, P.J.; Majcherczyk, A.; Kües, U.; von Bergen, M.; Buscot, F.;
Journal / Serie Enzyme and Microbial Technology
Erscheinungsjahr 2007
Department PROTEOM; BOOEK;
Band/Volume 41
Heft 6-7
Sprache englisch;
Keywords Copper binding region; Basidiomycetes; Native-PAGE; SDS-PAGE; Western blot; Tandem mass spectrometry
Abstract This study presents a combined method to analyze extracellular fungal laccases using a new anti-laccase antibody together with the identification of tryptic laccase peptides by mass spectrometry (nanoLC-ESI-MS/MS). The polyclonal anti-laccase antibody LccCbr2 was raised against peptides designed from the copper binding region II of fungal laccases using in silico data obtained from GenBank database. As a consequence, detection requires denaturation of the enzymes due to the stable conformation of the copper binding region II. The specificity of the antibody was shown with denatured laccase Lcc1 of Coprinopsis cinerea and laccase of Hypholoma fasciculare. LccCbr2 detected amounts as low as 5 ng of highly purified laccase, indicating a possible use of the antibody for quantification of laccase proteins. Denatured extracellular laccases from culture supernatants of the basidiomycetes C. cinerea, H. fasciculare, Lentinula edodes, Mycena sp., Piriformospora indica, Pleurotus cornucopiae, Pleurotus ostreatus, Pycnoporus cinnabarinus, Trametes versicolor and furthermore the ascomycete Verpa conica were detected with apparent molecular masses between 60 and 70 kDa by LccCbr2. The identity of extracellular laccases from C. cinerea, H. fasciculare, P. ostreatus, P. cinnabarinus and T. versicolor were verified by tryptic peptides using nanoLC-ESI-MS/MS.
ID 1922
dauerhafte UFZ-Verlinkung https://www.ufz.de/index.php?en=20939&ufzPublicationIdentifier=1922
Kellner, H., Jehmlich, N., Benndorf, D., Hoffmann, R., Rühl, M., Hoegger, P.J., Majcherczyk, A., Kües, U., von Bergen, M., Buscot, F. (2007):
Detection, quantification and identification of fungal extracellular laccases using polyclonal antibody and mass spectrometry
Enzyme Microb. Technol. 41 (6-7), 694 - 701