Endo-β-N-acetylglucosamidases (ENGases) in the fungus Trichoderma atroviride: possible involvement of the filamentous fungi-specific cytosolic ENGase in the ERAD process.
N-Glycosylation is an important post-translational modification of proteins, which mainly occurs in the endoplasmic reticulum (ER). Glycoproteins that are unable to fold properly are exported to the cytosol for degradation by a cellular system called ER-associated degradation (ERAD). Once misfolded glycoproteins are exported to the cytosol, they are subjected to deglycosylation by peptide:N-glycanase (PNGase) to facilitate the efficient degradation of misfolded proteins by the proteasome. Interestingly, the ortholog of PNGase in some filamentous fungi was found to be an inactive deglycosylating enzyme. On the other hand, it has been shown that in filamentous fungi genomes, usually two different fungi-specific endo-β-N-acetylglucosamidases (ENGases) can be found; one is predicted to be localized in the cytosol and the other to have a signal sequence, while the functional importance of these enzymes remains to be clarified. In this study the ENGases of the filamentous fungus Trichoderma atroviride was characterized. By heterologous expression of the ENGases Eng18A and Eng18B in Saccharomyces cerevisiae, it was found that both ENGases are active deglycosylating enzymes. Interestingly, only Eng18B was able to enhance the efficient degradation of the RTL protein, a PNGase-dependent ERAD substrate, implying the involvement of this enzyme in the ERAD process. These results indicate that T. atroviride Eng18B may deglycosylate misfolded glycoproteins, substituting the function of the cytoplasmic PNGase in the ERAD process.
Department of Forest Mycology and Plant Pathology, Uppsala Biocenter, Swedish University of Agricultural Sciences, Box 7026, 75007 Uppsala, Sweden. Electronic address: georgios.tzelepis@slu.se.
Georgios
G
Tzelepis
Glycometabolome Team, Systems Glycobiology Research Group, RIKEN-Max Planck Joint Research Center, RIKEN Global Research Cluster, 2-1 Hirosawa, Wako Saitama 351-0198, Japan.
Akira
A
Hosomi
Glycometabolome Team, Systems Glycobiology Research Group, RIKEN-Max Planck Joint Research Center, RIKEN Global Research Cluster, 2-1 Hirosawa, Wako Saitama 351-0198, Japan; Graduate School of Science and Engineering, Saitama University, 255 Shimo-Ohkubo, Sakura-ku, Saitama City, Saitama 338-8570, Japan.
Tanim Jabid
TJ
Hossain
Glycometabolome Team, Systems Glycobiology Research Group, RIKEN-Max Planck Joint Research Center, RIKEN Global Research Cluster, 2-1 Hirosawa, Wako Saitama 351-0198, Japan.
Hiroto
H
Hirayama
Department of Forest Mycology and Plant Pathology, Uppsala Biocenter, Swedish University of Agricultural Sciences, Box 7026, 75007 Uppsala, Sweden.
Mukesh
M
Dubey
Department of Forest Mycology and Plant Pathology, Uppsala Biocenter, Swedish University of Agricultural Sciences, Box 7026, 75007 Uppsala, Sweden.
Dan Funck
DF
Jensen
Glycometabolome Team, Systems Glycobiology Research Group, RIKEN-Max Planck Joint Research Center, RIKEN Global Research Cluster, 2-1 Hirosawa, Wako Saitama 351-0198, Japan; Graduate School of Science and Engineering, Saitama University, 255 Shimo-Ohkubo, Sakura-ku, Saitama City, Saitama 338-8570, Japan.
Tadashi
T
Suzuki
Department of Forest Mycology and Plant Pathology, Uppsala Biocenter, Swedish University of Agricultural Sciences, Box 7026, 75007 Uppsala, Sweden.
Magnus
M
Karlsson
D000118:Acetylglucosaminidase / Q000235:genetics / Q000378:metabolism*; D000595:Amino Acid Sequence; D003600:Cytosol / Q000378:metabolism; D060746:Endoplasmic Reticulum-Associated Degradation / Q000235:genetics; D005656:Fungal Proteins / Q000235:genetics / Q000378:metabolism*; D016681:Genome, Fungal; D006031:Glycosylation; D008969:Molecular Sequence Data; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000235:genetics / Q000378:metabolism; D017386:Sequence Homology, Amino Acid; D014242:Trichoderma / Q000235:genetics / Q000378:metabolism*
D005656:Fungal Proteins; D011994:Recombinant Proteins; D000118:Acetylglucosaminidase
De-N-glycosylation; GH18; Heterologous expression; PNGases