Expression of a catalytic domain of a Neocallimastix frontalis endoxylanase gene (xyn3) in Kluyveromyces lactis and Penicillium roqueforti.
A cDNA fragment encoding the A catalytic domain of the Neocallimastix frontalis endoxylanase XYN3 was amplified and cloned by the polymerase chain reaction technique. The xyn3A DNA fragment was inserted between the Saccharomyces cerevisiae phosphoglycerate kinase gene promoter and terminator sequences on a multicopy episomal plasmid for Kluyveromyces lactis. The XYN3A domain was successfully expressed in K. lactis and functional endoxylanase was secreted by the yeast cells with the K. lactis killer toxin secretion signal. The XYN3A domain was also expressed in a strain of Penicillium roqueforti as a fusion protein (ShBLE::XYN3A) of the phleomycin-resistance gene product and the endoxylanase. Active endoxylanase was efficiently secreted from the fungal cells with the Trichoderma viride cellobiohydrolase (CBH1) secretion signal and processed by a related KEX2 endoprotease of the secretion pathway. Several differently glycosylated forms of the recombinant enzymes were secreted by the yeast and the filamentous fungus.
Durand
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Laboratoire de Biologie Cellulaire Fongique, CNRS UMR 5534, Université Lyon I, Villeurbanne, France. rdurand@univ-littoral.fr
Rascle
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Fèvre
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D000595:Amino Acid Sequence; D001483:Base Sequence; D020134:Catalytic Domain / Q000235:genetics; D043364:Endo-1,4-beta Xylanases; D005800:Genes, Fungal; D007716:Kluyveromyces / Q000235:genetics*; D008969:Molecular Sequence Data; D020088:Neocallimastix / Q000201:enzymology / Q000235:genetics*; D010407:Penicillium / Q000235:genetics*; D010446:Peptide Fragments / Q000096:biosynthesis / Q000235:genetics; D010735:Phosphoglycerate Kinase / Q000235:genetics; D011401:Promoter Regions, Genetic; D011994:Recombinant Proteins / Q000096:biosynthesis*; D013728:Terminator Regions, Genetic; D014242:Trichoderma / Q000235:genetics; D014995:Xylosidases / Q000096:biosynthesis* / Q000235:genetics
D010446:Peptide Fragments; D011994:Recombinant Proteins; D010735:Phosphoglycerate Kinase; D014995:Xylosidases; D043364:Endo-1,4-beta Xylanases