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Glycan remodeling with processing inhibitors and lectin-resistant eukaryotic cells

Glycan remodeling with processing inhibitors and lectin-resistant eukaryotic cells
Glycan remodeling with processing inhibitors and lectin-resistant eukaryotic cells

Some of the most important and interesting molecules in metazoan biology are glycoproteins. The importance of the carbohydrate component of these structures is often revealed by the disease phenotypes that manifest when the biosynthesis of particular glycoforms is disrupted. On the other hand, the presence of large amounts of carbohydrate can often hinder the structural and functional analysis of glycoproteins. There are often good reasons, therefore, for wanting to engineer and predefine the N -glycans present on glycoproteins, e.g., in order to characterize the functions of the glycans or facilitate their subsequent removal. Here, we describe in detail two distinct ways in which to usefully interfere with oligosaccharide processing, one involving the use of specific processing inhibitors, and the other the selection of cell lines mutated at gene loci that control oligosaccharide processing, using cytotoxic lectins. Both approaches have the capacity for controlled, radical alteration of oligosaccharide processing in eukaryotic cells used for heterologous protein expression, and have great utility in the structural analysis of glycoproteins.

Kifunensine, Lectin selection, Mammalian expression, N-glycan processing, Protein crystallization
307-322
Humana Press
Chang, Veronica T.
e83571e9-6b2f-4f8e-adfc-3fd539e9102e
Spooner, Robert A.
f8b5e206-ebd5-4d69-a256-b3195025fa19
Crispin, Max
cd980957-0943-4b89-b2b2-710f01f33bc9
Davis, Simon J.
77c9e91c-a2c6-498a-9128-bf164a8da8de
Castilho, A.
Chang, Veronica T.
e83571e9-6b2f-4f8e-adfc-3fd539e9102e
Spooner, Robert A.
f8b5e206-ebd5-4d69-a256-b3195025fa19
Crispin, Max
cd980957-0943-4b89-b2b2-710f01f33bc9
Davis, Simon J.
77c9e91c-a2c6-498a-9128-bf164a8da8de
Castilho, A.

Chang, Veronica T., Spooner, Robert A., Crispin, Max and Davis, Simon J. (2015) Glycan remodeling with processing inhibitors and lectin-resistant eukaryotic cells. In, Castilho, A. (ed.) Glyco-Engineering. (Methods in Molecular Biology, , (doi:10.1007/978-1-4939-2760-9_21), 1321) New York, NY. Humana Press, pp. 307-322. (doi:10.1007/978-1-4939-2760-9_21).

Record type: Book Section

Abstract

Some of the most important and interesting molecules in metazoan biology are glycoproteins. The importance of the carbohydrate component of these structures is often revealed by the disease phenotypes that manifest when the biosynthesis of particular glycoforms is disrupted. On the other hand, the presence of large amounts of carbohydrate can often hinder the structural and functional analysis of glycoproteins. There are often good reasons, therefore, for wanting to engineer and predefine the N -glycans present on glycoproteins, e.g., in order to characterize the functions of the glycans or facilitate their subsequent removal. Here, we describe in detail two distinct ways in which to usefully interfere with oligosaccharide processing, one involving the use of specific processing inhibitors, and the other the selection of cell lines mutated at gene loci that control oligosaccharide processing, using cytotoxic lectins. Both approaches have the capacity for controlled, radical alteration of oligosaccharide processing in eukaryotic cells used for heterologous protein expression, and have great utility in the structural analysis of glycoproteins.

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More information

Published date: 16 June 2015
Keywords: Kifunensine, Lectin selection, Mammalian expression, N-glycan processing, Protein crystallization

Identifiers

Local EPrints ID: 414314
URI: https://eprints.soton.ac.uk/id/eprint/414314
PURE UUID: 95b6ebb1-da3f-40de-930c-1bffb9a3ccda
ORCID for Max Crispin: ORCID iD orcid.org/0000-0002-1072-2694

Catalogue record

Date deposited: 26 Sep 2017 16:30
Last modified: 14 Mar 2019 01:25

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