title: The Role of Conserved N-Linked Glycans on Ebola Virus Glycoprotein 2.
abstract: BACKGROUND
  N-linked glycosylation is a common posttranslational modification found on viral glycoproteins (GPs) and involved in promoting expression, cellular attachment, protection from proteases, and antibody evasion. The GP subunit GP2 of filoviruses contains 2 completely conserved N-linked glycosylation sites (NGSs) at N563 and N618, suggesting that they have been maintained through selective pressures.
  
  
  METHODS
  We assessed mutants lacking these glycans for expression and function to understand the role of these sites during Ebola virus entry.
  
  
  RESULTS
  Elimination of either GP2 glycan individually had a modest effect on GP expression and no impact on antibody neutralization of vesicular stomatitis virus pseudotyped with Ebola virus GP. However, loss of the N563 glycan enhanced entry by 2-fold and eliminated GP detection by a well-characterized monoclonal antibody KZ52. Loss of both sites dramatically decreased GP expression and abolished entry. Surprisingly, a GP that retained a single NGS at N563, eliminating the remaining 16 NGSs from GP1 and GP2, had detectable expression, a modest increase in entry, and pronounced sensitivity to antibody neutralization.
  
  
  CONCLUSIONS
  Our findings support the importance of the GP2 glycans in GP expression/structure, transduction efficiency, and antibody neutralization, particularly when N-linked glycans are also removed from GP1.
authors.affiliation: Department of Microbiology, University of Iowa, Iowa City.
authors.forename: Nicholas J
authors.initials: NJ
authors.lastname: Lennemann
authors.affiliation: Department of Microbiology, University of Iowa, Iowa City.
authors.forename: Madeline
authors.initials: M
authors.lastname: Walkner
authors.affiliation: Department of Microbiology, University of Iowa, Iowa City.
authors.forename: Abigail R
authors.initials: AR
authors.lastname: Berkebile
authors.affiliation: Department of Microbiology, University of Iowa, Iowa City.
authors.forename: Neil
authors.initials: N
authors.lastname: Patel
authors.affiliation: Department of Microbiology, University of Iowa, Iowa City.
authors.forename: Wendy
authors.initials: W
authors.lastname: Maury
mesh_terms: D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology; D057134:Antibodies, Neutralizing / Q000276:immunology; D002460:Cell Line; D002522:Chlorocebus aethiops; D017124:Conserved Sequence / Q000235:genetics*; D029043:Ebolavirus / Q000235:genetics* / Q000276:immunology / Q000472:pathogenicity*; D057809:HEK293 Cells; D019142:Hemorrhagic Fever, Ebola / Q000276:immunology / Q000821:virology*; D006801:Humans; D009154:Mutation / Q000235:genetics; D011134:Polysaccharides / Q000235:genetics*; D014709:Vero Cells; D018116:Vesiculovirus / Q000276:immunology; D014759:Viral Envelope Proteins / Q000235:genetics*; D053586:Virus Internalization
chemical_list: D000911:Antibodies, Monoclonal; D057134:Antibodies, Neutralizing; D011134:Polysaccharides; D014759:Viral Envelope Proteins; C113189:envelope glycoprotein, Ebola virus
keywords: Ebola virus; N-linked glycosylation; antibody neutralization; filovirus; glycan; glycoprotein; mutagenesis; virus entry