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PMID: 17355862 Published · ppublish English Journal Article Research Support, Non-U.S. Gov't

Glycoprotein structural genomics: solving the glycosylation problem.

Structure (London, England : 1993) ·Vol. 15 ·No. 3 ·2007-03-00 ·Pages 267-73

Chang VT, Crispin M, Aricescu AR, Harvey DJ, Nettleship JE, Fennelly JA, Yu C, Boles KS, Evans EJ, Stuart DI, Dwek RA, Jones EY, Owens RJ, Davis SJ

Abstract

Glycoproteins present special problems for structural genomic analysis because they often require glycosylation in order to fold correctly, whereas their chemical and conformational heterogeneity generally inhibits crystallization. We show that the "glycosylation problem" can be solved by expressing glycoproteins transiently in mammalian cells in the presence of the N-glycosylation processing inhibitors, kifunensine or swainsonine. This allows the correct folding of the glycoproteins, but leaves them sensitive to enzymes, such as endoglycosidase H, that reduce the N-glycans to single residues, enhancing crystallization. Since the scalability of transient mammalian expression is now comparable to that of bacterial systems, this approach should relieve one of the major bottlenecks in structural genomic analysis.

MeSH Terms
Alkaloids/chemistry,genetics,metabolism Cell Line Cloning, Molecular Enzyme Inhibitors Genomics/methods Glycoproteins/antagonists & inhibitors,chemistry,genetics,metabolism Glycosylation/drug effects Humans Swainsonine/chemistry,metabolism
Chemicals
Alkaloids Enzyme Inhibitors Glycoproteins kifunensine Swainsonine
Authors & Affiliations
14 authors, click to expand affiliations / ORCID
Chang Veronica T
Nuffield Department of Clinical Medicine and MRC Human Immunology Unit, Weatherall Institute of Molecular Medicine, University of Oxford, Oxford OX3 9DS, United Kingdom.
Crispin Max
Aricescu A Radu
Harvey David J
Nettleship Joanne E
Fennelly Janet A
Yu Chao
Boles Kent S
Evans Edward J
Stuart David I
Dwek Raymond A
Jones E Yvonne
Owens Raymond J
Davis Simon J
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Article Info
Journal
Structure (London, England : 1993)
Abbr.
Structure
ISSN
0969-2126
DOI
10.1016/j.str.2007.01.011
Published
2007-03-00
Pages
267-73
Language
English
Region
United States
NLM ID
101087697
PMCID
PMC1885966
Subset
IM
Grants
Wellcome Trust · 081894 · United Kingdom
Medical Research Council · G0400717 · United Kingdom
Medical Research Council · G9900061 · United Kingdom
External Links
PubMed source DOI: 10.1016/j.str.2007.01.011 PMC Full Text View journal details
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