BACKGROUND: The cytoplasmic peptide:N-glycanase (PNGase) is a deglycosylating enzyme involved in the ER-associated degradation (ERAD) process, while ERAD-independent activities are also reported. Previous biochemical analyses indicated that the cytoplasmic PNGase orthologue in Arabidopsis thaliana (AtPNG1) can function as not only PNGase but also transglutaminase, while its in vivo function remained unclarified. METHODS: AtPNG1 was expressed in Saccharomyces cerevisiae and its in vivo role on PNGase-dependent ERAD pathway was examined. RESULTS: AtPNG1 could facilitate the ERAD through its deglycosylation activity. Moreover, a catalytic mutant of AtPNG1 (AtPNG1(C251A)) was found to significantly impair the ERAD process. This result was found to be N-glycan-dependent, as the AtPNG(C251A) did not affect the stability of the non-glycosylated RTA∆ (ricin A chain non-toxic mutant). Tight interaction between AtPNG1(C251A) and the RTA∆ was confirmed by co-immunoprecipitation analysis. CONCLUSION: The plant PNGase facilitates ERAD through its deglycosylation activity, while the catalytic mutant of AtPNG1 impair glycoprotein ERAD by binding to N-glycans on the ERAD substrates. GENERAL SIGNIFICANCE: Our studies underscore the functional importance of a plant PNGase orthologue as a deglycosylating enzyme involved in the ERAD. Crown
BACKGROUND: The cytoplasmic peptide:N-glycanase (PNGase) is a deglycosylating enzyme involved in the ER-associated degradation (ERAD) process, while ERAD-independent activities are also reported. Previous biochemical analyses indicated that the cytoplasmic PNGase orthologue in Arabidopsis thaliana (AtPNG1) can function as not only PNGase but also transglutaminase, while its in vivo function remained unclarified. METHODS:AtPNG1 was expressed in Saccharomyces cerevisiae and its in vivo role on PNGase-dependent ERAD pathway was examined. RESULTS:AtPNG1 could facilitate the ERAD through its deglycosylation activity. Moreover, a catalytic mutant of AtPNG1 (AtPNG1(C251A)) was found to significantly impair the ERAD process. This result was found to be N-glycan-dependent, as the AtPNG(C251A) did not affect the stability of the non-glycosylated RTA∆ (ricin A chain non-toxic mutant). Tight interaction between AtPNG1(C251A) and the RTA∆ was confirmed by co-immunoprecipitation analysis. CONCLUSION: The plant PNGase facilitates ERAD through its deglycosylation activity, while the catalytic mutant of AtPNG1 impair glycoprotein ERAD by binding to N-glycans on the ERAD substrates. GENERAL SIGNIFICANCE: Our studies underscore the functional importance of a plant PNGase orthologue as a deglycosylating enzyme involved in the ERAD. Crown
Authors: Ping He; Jeff E Grotzke; Bobby G Ng; Murat Gunel; Hamed Jafar-Nejad; Peter Cresswell; Gregory M Enns; Hudson H Freeze Journal: Glycobiology Date: 2015-04-21 Impact factor: 4.313
Authors: Gregory M Enns; Vandana Shashi; Matthew Bainbridge; Michael J Gambello; Farah R Zahir; Thomas Bast; Rebecca Crimian; Kelly Schoch; Julia Platt; Rachel Cox; Jonathan A Bernstein; Mena Scavina; Rhonda S Walter; Audrey Bibb; Melanie Jones; Madhuri Hegde; Brett H Graham; Anna C Need; Angelica Oviedo; Christian P Schaaf; Sean Boyle; Atul J Butte; Rui Chen; Rong Chen; Michael J Clark; Rajini Haraksingh; Tina M Cowan; Ping He; Sylvie Langlois; Huda Y Zoghbi; Michael Snyder; Richard A Gibbs; Hudson H Freeze; David B Goldstein Journal: Genet Med Date: 2014-03-20 Impact factor: 8.822