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Title: Identification of functional elements of the GDP-fucose transporter SLC35C1 using a novel Chinese hamster ovary mutant
Authors: Zhang, P.
Haryadi, R.
Chan, K.F.
Teo, G.
Goh, J.
Pereira, N.A.
Feng, H.
Song, Z. 
Keywords: GDP-fucose transporter
Glycosylation mutant
Protein fucosylation
Structure-function relationships
Zinc-finger nucleases
Issue Date: Jul-2012
Citation: Zhang, P., Haryadi, R., Chan, K.F., Teo, G., Goh, J., Pereira, N.A., Feng, H., Song, Z. (2012-07). Identification of functional elements of the GDP-fucose transporter SLC35C1 using a novel Chinese hamster ovary mutant. Glycobiology 22 (7) : 897-911. ScholarBank@NUS Repository.
Abstract: The GDP-fucose transporter SLC35C1 critically regulates the fucosylation of glycans. Elucidation of its structure-function relationships remains a challenge due to the lack of an appropriate mutant cell line. Here we report a novel Chinese hamster ovary (CHO) mutant, CHO-gmt5, generated by the zinc-finger nuclease technology, in which the Slc35c1 gene was knocked out from a previously reported CHO mutant that has a dysfunctional CMP-sialic acid transporter (CST) gene (Slc35a1). Consequently, CHO-gmt5 harbors double genetic defects in Slc35a1 and Slc35c1 and produces N-glycans deficient in both sialic acid and fucose. The structure-function relationships of SLC35C1 were studied using CHO-gmt5 cells. In contrast to the CST and UDP-galactose transporter, the C-terminal tail of SLC35C1 is not required for its Golgi localization but is essential for generating glycans that are recognized by a fucose-binding lectin, Aleuria aurantia lectin (AAL), suggesting an important role in the transport activity of SLC35C1. Furthermore, we found that this impact can be independently contributed by a cluster of three lysine residues and a Glu-Met (EM) sequence within the C terminus. We also showed that the conserved glycine residues at positions 180 and 277 of SLC35C1 have significant impacts on AAL binding to CHO-gmt5 cells, suggesting that these conserved glycine residues are required for the transport activity of Slc35 proteins. The absence of sialic acid and fucose on Fc N-glycan has been independently shown to enhance the antibody-dependent cellular cytotoxicity (ADCC) effect. By combining these features into one cell line, we postulate that CHO-gmt5 may represent a more advantageous cell line for the production of recombinant antibodies with enhanced ADCC effect. © 2012 The Author.
Source Title: Glycobiology
ISSN: 09596658
DOI: 10.1093/glycob/cws064
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