Comparison of modular and non-modular xylanases as carrier proteins for the efficient secretion of heterologous proteins from Penicillium funiculosum.
Genes encoding three enzymes with xylanase activity from the filamentous fungus Penicillium funiculosum are described. Two of the encoded xylanases are predicted to be modular in structure with catalytic and substrate-binding domains separated by a serine and threonine-rich linker region; the other had none of these properties and was non-modular. In order to develop P. funiculosum as a host for the secreted production of heterologous proteins, each of the xylanases was assessed for use as a carrier protein in a fusion strategy. We show that one of the modular xylanases (encoded by xynA) was an effective carrier protein but the other (encoded by xynB) and the non-modular xylanase (encoded by xynC) were not effective as secretion carriers. We show that the beta-glucuronidase (GUS) protein from Escherichia coli is secreted by P. funiculosum when expressed as an XYNA fusion but that the secreted GUS protein, cleaved in vivo from XYNA, is glycosylated and enzymatically inactive.
Alcocer
M J C
MJ
School of Life and Environmental Sciences, University of Nottingham, University Park, NG7 2RD, Nottingham, UK.marcos.alcocer@nottingham.ac.uk
Furniss
C S M
CS
Kroon
P A
PA
Campbell
M
M
Archer
D B
DB
D000595:Amino Acid Sequence; D001665:Binding Sites; D002352:Carrier Proteins / Q000378:metabolism*; D020134:Catalytic Domain; D003001:Cloning, Molecular; D043364:Endo-1,4-beta Xylanases; D004926:Escherichia coli / Q000201:enzymology / Q000235:genetics; D005656:Fungal Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D005800:Genes, Fungal; D005813:Genes, Synthetic; D005966:Glucuronidase / Q000378:metabolism; D006031:Glycosylation; D006657:Histones / Q000235:genetics; D007527:Isoenzymes / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D008969:Molecular Sequence Data; D010407:Penicillium / Q000201:enzymology* / Q000235:genetics; D011401:Promoter Regions, Genetic; D011499:Protein Processing, Post-Translational; D017434:Protein Structure, Tertiary; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism*; D017386:Sequence Homology, Amino Acid; D014170:Transformation, Genetic; D043325:Xylan Endo-1,3-beta-Xylosidase; D014995:Xylosidases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D001617:beta-Glucosidase / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*
D002352:Carrier Proteins; D005656:Fungal Proteins; D006657:Histones; D007527:Isoenzymes; D011993:Recombinant Fusion Proteins; D014995:Xylosidases; D001617:beta-Glucosidase; D005966:Glucuronidase; D043325:Xylan Endo-1,3-beta-Xylosidase; D043364:Endo-1,4-beta Xylanases; C090001:XynB xylanase