Ghent University Academic Bibliography

Advanced

High resolution crystal structure of the endo-N-acetyl-β-D-glucosaminidase responsible for the deglycosylation of Hypocrea jecorina cellulases

Ingeborg Stals UGent, Saeid Karkehabadi, Steve Kim, Michael Ward, Anita Van Landschoot UGent, Bart Devreese UGent and Mats Sandgren (2012) PLOS ONE. 7(7).
abstract
Endo-N-acetyl-β-D-glucosaminidases hydrolyze the glycosidic linkage between the two N-acetylglucosamine units that make up the chitobiose core of N-glycans. The endo-N- acetyl-β-D-glucosaminidases classified into glycoside hydrolase family 18 are small, bacterial proteins with different substrate specificities. Recently two eukaryotic family 18 deglycosylating enzymes have been identified. Here, the expression, purification and the 1.3Å resolution structure of the ENGase (Endo T) from the mesophilic fungus Hypocrea jecorina (anamorph Trichoderma reesei) are reported. Although the mature protein is C-terminally processed with a 43 amino acid peptide, the protein has a complete (β/α)8 Tim-barrel topology. In the active site, the proton donor (E131) and the residue stabilizing the transition state (D129) in the substrate assisted catalysis mechanism are found in almost identical positions as in the bacterial GH18 ENGases: Endo H, Endo F1, Endo F3 and Endo BT. However, the loops defining the substrate- binding cleft have great variation compared to the previously known ENGase structures, and the structures also differ in some of the α-helices forming the barrel. This could reflect the variation in substrate specificity between the five enzymes. This is the first three dimensional structure of a eukaryotic endo-N-acetyl-β-D- glucosaminidase from glycoside hydrolase family 18. A glycosylation analysis of the cellulases secreted by a Hypocrea jecorina Endo T knock-out strain shows the in vivo function of the protein. A homology search and phylogenetic analysis show that the two known enzymes and their homologues form a large but separate cluster in subgroup B of the fungal chitinases. Therefore the future use of a uniform nomenclature is proposed.
Please use this url to cite or link to this publication:
author
organization
alternative title
High resolution crystal structure of the endo-N-acetyl-beta-D-glucosaminidase responsible for the deglycosylation of Hypocrea jecorina cellulases
year
type
journalArticle (original)
publication status
published
subject
keyword
protein structure, EC 3.2.1.96, Hypocrea jecorina, endo-β-N-acetylglucosaminidase, glycoside hydrolase family 18, cellulases, deglycosylation, knock-out strain, overexpression
journal title
PLOS ONE
PLoS One
volume
7
issue
7
article_number
e40854
pages
13 pages
Web of Science type
Article
Web of Science id
000306950900013
JCR category
MULTIDISCIPLINARY SCIENCES
JCR impact factor
3.73 (2012)
JCR rank
7/56 (2012)
JCR quartile
1 (2012)
ISSN
1932-6203
DOI
10.1371/journal.pone.0040854
project
OF - Enzymatische technologie voor industriële processen
language
English
UGent publication?
yes
classification
A1
copyright statement
I have retained and own the full copyright for this publication
id
2124025
handle
http://hdl.handle.net/1854/LU-2124025
date created
2012-05-31 09:48:25
date last changed
2013-07-15 14:12:55
@article{2124025,
  abstract     = {Endo-N-acetyl-\ensuremath{\beta}-D-glucosaminidases hydrolyze the glycosidic linkage between the two N-acetylglucosamine units that make up the chitobiose core of N-glycans. The endo-N- acetyl-\ensuremath{\beta}-D-glucosaminidases classified into glycoside hydrolase family 18 are small, bacterial proteins with different substrate specificities. Recently two eukaryotic family 18 deglycosylating enzymes have been identified. Here, the expression, purification and the 1.3A\unmatched{030a} resolution structure of the ENGase (Endo T) from the mesophilic fungus Hypocrea jecorina (anamorph Trichoderma reesei) are reported. Although the mature protein is C-terminally processed with a 43 amino acid peptide, the protein has a complete (\ensuremath{\beta}/\ensuremath{\alpha})8 Tim-barrel topology. In the active site, the proton donor (E131) and the residue stabilizing the transition state (D129) in the substrate assisted catalysis mechanism are found in almost identical positions as in the bacterial GH18 ENGases: Endo H, Endo F1, Endo F3 and Endo BT. However, the loops defining the substrate- binding cleft have great variation compared to the previously known ENGase structures, and the structures also differ in some of the \ensuremath{\alpha}-helices forming the barrel. This could reflect the variation in substrate specificity between the five enzymes. This is the first three dimensional structure of a eukaryotic endo-N-acetyl-\ensuremath{\beta}-D- glucosaminidase from glycoside hydrolase family 18. A glycosylation analysis of the cellulases secreted by a Hypocrea jecorina Endo T knock-out strain shows the in vivo function of the protein. A homology search and phylogenetic analysis show that the two known enzymes and their homologues form a large but separate cluster in subgroup B of the fungal chitinases. Therefore the future use of a uniform nomenclature is proposed.},
  articleno    = {e40854},
  author       = {Stals, Ingeborg and Karkehabadi, Saeid  and Kim, Steve and Ward, Michael and Van Landschoot, Anita and Devreese, Bart and Sandgren, Mats},
  issn         = {1932-6203},
  journal      = {PLOS ONE},
  keyword      = {protein structure,EC 3.2.1.96,Hypocrea jecorina,endo-\ensuremath{\beta}-N-acetylglucosaminidase,glycoside hydrolase family 18,cellulases,deglycosylation,knock-out strain,overexpression},
  language     = {eng},
  number       = {7},
  pages        = {13},
  title        = {High resolution crystal structure of the endo-N-acetyl-\ensuremath{\beta}-D-glucosaminidase responsible for the deglycosylation of Hypocrea jecorina cellulases},
  url          = {http://dx.doi.org/10.1371/journal.pone.0040854},
  volume       = {7},
  year         = {2012},
}

Chicago
Stals, Ingeborg, Saeid Karkehabadi, Steve Kim, Michael Ward, Anita Van Landschoot, Bart Devreese, and Mats Sandgren. 2012. “High Resolution Crystal Structure of the endo-N-acetyl-β-D-glucosaminidase Responsible for the Deglycosylation of Hypocrea Jecorina Cellulases.” Plos One 7 (7).
APA
Stals, I., Karkehabadi, S., Kim, S., Ward, M., Van Landschoot, A., Devreese, B., & Sandgren, M. (2012). High resolution crystal structure of the endo-N-acetyl-β-D-glucosaminidase responsible for the deglycosylation of Hypocrea jecorina cellulases. PLOS ONE, 7(7).
Vancouver
1.
Stals I, Karkehabadi S, Kim S, Ward M, Van Landschoot A, Devreese B, et al. High resolution crystal structure of the endo-N-acetyl-β-D-glucosaminidase responsible for the deglycosylation of Hypocrea jecorina cellulases. PLOS ONE. 2012;7(7).
MLA
Stals, Ingeborg, Saeid Karkehabadi, Steve Kim, et al. “High Resolution Crystal Structure of the endo-N-acetyl-β-D-glucosaminidase Responsible for the Deglycosylation of Hypocrea Jecorina Cellulases.” PLOS ONE 7.7 (2012): n. pag. Print.