Characterization of glyco isoforms in plasma-derived human antithrombin by on-line capillary zone electrophoresis-electrospray ionization-quadrupole ion trap-mass spectrometry of the intact glycoproteins.
The carbohydrate structures of five isoforms of alpha-AT and two isoforms of beta-AT were determined by applying capillary zone electrophoresis (CZE) on-line coupled to electrospray ionization-mass spectrometry (ESI-MS) using an ion-trap analyzer. For the AT preparations gained from a plasma pool at least semiquantitative information on the isoform-distributions could be gained. Unlike to the commonly used approaches starting from enzymatically treated glycoproteins, this approach deals with intact proteins. The high accuracy of the molecular mass determination obtained by the ion-trap analyzer allows one to calculate and ascertain the carbohydrate composition assuming no variations in the protein moiety of AT and to exclude or confirm the presence of the potential post-translational or other modifications. Therefore, the direct coupling of CZE with ESI-MS does not only represent a fast alternative technique to two-dimensional electrophoresis (2-DE) but serves as a method which provides structural information complementary to that gained from peptide mapping methods.
Demelbauer
Uwe M
UM
Institute of Analytical Chemistry, University of Vienna.
Plematl
Alexander
A
Institute of Analytical Chemistry, University of Vienna.
Kremser
Leopold
L
Institute of Analytical Chemistry, University of Vienna.
Allmaier
Günter
G
Institute of Chemical Technologies and Analytics, Vienna University of Technology.
Josic
Djuro
D
Octapharma Pharmazeutika Produktions Gesellschaft, Vienna, Austria.
Rizzi
Andreas
A
Institute of Analytical Chemistry, University of Vienna.
D000991:Antithrombins / Q000032:analysis* / Q000737:chemistry; D002240:Carbohydrate Sequence; D002241:Carbohydrates / Q000737:chemistry; D019075:Electrophoresis, Capillary / Q000379:methods*; D006023:Glycoproteins / Q000032:analysis* / Q000737:chemistry; D006801:Humans; D008968:Molecular Conformation; D008969:Molecular Sequence Data; D010449:Peptide Mapping; D010949:Plasma / Q000737:chemistry*; D020033:Protein Isoforms / Q000032:analysis* / Q000737:chemistry; D011499:Protein Processing, Post-Translational; D021241:Spectrometry, Mass, Electrospray Ionization / Q000379:methods*; D014508:Urea / Q000737:chemistry
D000991:Antithrombins; D002241:Carbohydrates; D006023:Glycoproteins; D020033:Protein Isoforms; D014508:Urea