title: Plasma N-glycan profiling by mass spectrometry for congenital disorders of glycosylation type II.
abstract: BACKGROUND
  Determination of the genetic defect in patients with a congenital disorder of glycosylation (CDG) is challenging because of the wide clinical presentation, the large number of gene products involved, and the occurrence of secondary causes of underglycosylation. Transferrin isoelectric focusing has been the method of choice for CDG screening; however, improved methods are required for the molecular diagnosis of patients with CDG type II.
  
  
  METHODS
  Plasma samples with a typical transferrin isofocusing profile were analyzed. N-glycans were released from these samples by PNGase F [peptide-N4-(acetyl-β-glucosaminyl)-asparagine amidase] digestion, permethylated and purified, and measured on a MALDI linear ion trap mass spectrometer. A set of 38 glycans was used for quantitative comparison and to establish reference intervals for such glycan features as the number of antennae, the level of truncation, and fucosylation. Plasma N-glycans from control individuals, patients with known CDG type II defects, and patients with a secondary cause of underglycosylation were analyzed.
  
  
  RESULTS
  CDGs due to mannosyl (α-1,6-)-glycoprotein β-1,2-N-acetylglucosaminyltransferase (MGAT2), β-1,4-galactosyltransferase 1 (B4GALT1), and SLC35C1 (a GDP-fucose transporter) defects could be diagnosed directly from the N-glycan profile. CDGs due to defects in proteins involved in Golgi trafficking, such as subunit 7 of the conserved oligomeric Golgi complex (COG7) and subunit V0 a2 of the lysosomal H(+)-transporting ATPase (ATP6V0A2) caused a loss of triantennary N-glycans and an increase of truncated structures. Secondary causes with liver involvement were characterized by increased fucosylation, whereas the presence of plasma sialidase produced isolated undersialylation.
  
  
  CONCLUSIONS
  MALDI ion trap analysis of plasma N-glycans documents features that discriminate between primary and secondary causes of underglycosylation and should be applied as the first step in the diagnostic track of all patients with an unsolved CDG type II.
authors.affiliation: Department of Laboratory Medicine, Radboud University Nijmegen Medical Centre, Nijmegen, The Netherlands.
authors.forename: Maïlys
authors.initials: M
authors.lastname: Guillard
authors.forename: Eva
authors.initials: E
authors.lastname: Morava
authors.forename: Floris L
authors.initials: FL
authors.lastname: van Delft
authors.forename: Rosie
authors.initials: R
authors.lastname: Hague
authors.forename: Christian
authors.initials: C
authors.lastname: Körner
authors.forename: Maciej
authors.initials: M
authors.lastname: Adamowicz
authors.forename: Ron A
authors.initials: RA
authors.lastname: Wevers
authors.forename: Dirk J
authors.initials: DJ
authors.lastname: Lefeber
mesh_terms: D016022:Case-Control Studies; D018981:Congenital Disorders of Glycosylation / Q000097:blood / Q000175:diagnosis*; D006801:Humans; D007231:Infant, Newborn; D015997:Neonatal Screening; D011134:Polysaccharides / Q000097:blood*; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization / Q000379:methods*
chemical_list: D011134:Polysaccharides