Skip Navigation

This Article
Right arrow Full Text Freely available
Right arrow FREE Full Text (PDF) Freely available
Right arrow Alert me when this article is cited
Right arrow Alert me if a correction is posted
Services
Right arrow Email this article to a friend
Right arrow Similar articles in this journal
Right arrow Similar articles in ISI Web of Science
Right arrow Similar articles in PubMed
Right arrow Alert me to new issues of the journal
Right arrow Add to My Personal Archive
Right arrow Download to citation manager
Right arrow Search for citing articles in:
ISI Web of Science (24)
Right arrowRequest Permissions
Right arrow Disclaimer
Google Scholar
Right arrow Articles by Kitova, E. N.
Right arrow Articles by Klassen, J. S.
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Kitova, E. N.
Right arrow Articles by Klassen, J. S.
Social Bookmarking
Add to CiteULike   Add to Connotea   Add to Del.icio.us  
What's this?

Glycobiology, 2001, Vol. 11, No. 7 605-611
© 2001 Oxford University Press

The observation of multivalent complexes of Shiga-like toxin with globotriaoside and the determination of their stoichiometry by nanoelectrospray Fourier-transform ion cyclotron resonance mass spectrometry

Elena N. Kitova, Pavel I. Kitov, David R. Bundle and John S. Klassen1

Department of Chemistry, University of Alberta, Edmonton, Alberta, Canada T6G 2G2

We show by nanoelectrospray ionization (nanoES) Fourier-transform ion cyclotron resonance mass spectrometry (FT-ICR MS) that it is possible to observe oligosaccharide–protein complexes with dissociation constants in the millimolar range, such as Pk trisaccharide (globotriaoside) complexed with the Shiga-like toxin (SLT) of pathogenic E. coli. It is further demonstrated that nanoES/FT-ICR MS is an exquisite method to study quantitative aspects of the association of mono- and polyvalent oligosaccharide ligands with multimeric proteins, such as the SLTs. At increasing trisaccharide:protein ratios it was shown that the B5 toxin subunit complexes with 5 Pk trisaccharides and only after all 5 copies of site 2 are essentially filled do any of the remaining 10 receptor sites become occupied. From the distribution of bound Pk’s at the five binding sites, it was possible to establish association constants for each of the five sites and to confirm that binding occurs noncooperatively, the association constants for each site are identical and that compared to site 1, site 2 exhibits a tenfold higher affinity for the globotriaoside synthetic ligand 1. The facile identification of the occupancy of binding sites represents information that is not readily available by other techniques. This sensitive and rapid estimation of association constants for protein–ligand complexes, which are free of unpredictable secondary effects that plague enzyme linked assays, is likely to find wide application.

1 To whom correspondence should be addressed


Add to CiteULike CiteULike   Add to Connotea Connotea   Add to Del.icio.us Del.icio.us    What's this?


This article has been cited by other articles:


Home page
 GlycobiologyHome page
G. K Shoemaker, N. Soya, M. M Palcic, and J. S Klassen
Temperature-dependent cooperativity in donor-acceptor substrate binding to the human blood group glycosyltransferases
Glycobiology, August 1, 2008; 18(8): 587 - 592.
[Abstract] [Full Text] [PDF]

Home page
 GlycobiologyHome page
E. N Kitova, P. I Kitov, E. Paszkiewicz, J. Kim, G. L Mulvey, G. D Armstrong, D. R Bundle, and J. S Klassen
Affinities of Shiga toxins 1 and 2 for univalent and oligovalent Pk-trisaccharide analogs measured by electrospray ionization mass spectrometry
Glycobiology, October 1, 2007; 17(10): 1127 - 1137.
[Abstract] [Full Text] [PDF]

Home page
 J. Biol. Chem.Home page
J. G. S. Ho, P. I. Kitov, E. Paszkiewicz, J. Sadowska, D. R. Bundle, and K. K.-S. Ng
Ligand-assisted Aggregation of Proteins: DIMERIZATION OF SERUM AMYLOID P COMPONENT BY BIVALENT LIGANDS
J. Biol. Chem., September 9, 2005; 280(36): 31999 - 32008.
[Abstract] [Full Text] [PDF]

Home page
 J. Biol. Chem.Home page
A. M. Soltyk, C. R. MacKenzie, V. M. Wolski, T. Hirama, P. I. Kitov, D. R. Bundle, and J. L. Brunton
A Mutational Analysis of the Globotriaosylceramide-binding Sites of Verotoxin VT1
J. Biol. Chem., February 8, 2002; 277(7): 5351 - 5359.
[Abstract] [Full Text] [PDF]


Disclaimer: Please note that abstracts for content published before 1996 were created through digital scanning and may therefore not exactly replicate the text of the original print issues. All efforts have been made to ensure accuracy, but the Publisher will not be held responsible for any remaining inaccuracies. If you require any further clarification, please contact our Customer Services Department.