The Golgi CMP-sialic acid transporter: A new CHO mutant provides functional insights.

Glycobiology
Sing Fee LimZhiwei Song

Abstract

A CHO mutant line, MAR-11, was isolated using a cytotoxic lectin, Maackia amurensis agglutinin (MAA). This mutant has decreased levels of cell surface sialic acid relative to both wild-type CHO-K1 and Lec2 mutant CHO cells. The CMP-sialic acid transporter (CMP-SAT) gene in the MAR-11 mutant cell has a C-T mutation that results in a premature stop codon. As a result, MAR-11 cells express a truncated version of CMP-SAT which contains only 100 amino acids rather than the normal CMP-SAT which contains 336 amino acids. Biochemical analyses indicate that recombinant interferon-gamma (IFN-gamma) produced by the mutant cells lack sialic acid. Using MAR-11 as host cells, an EPO/IEF assay for the structure-function study of CMP-SAT was developed. This assay seems more sensitive than previous assays that were used to analyze sialylation in Lec2 cells. Cotransfection of constructs that express CMP-SAT into MAR-11 cells completely converted the recombinant EPO to a sialylation pattern that is similar to the EPO produced by the wild-type CHO cells. Using this assay, we showed that CMP-SAT lacking C-terminal 18 amino acids from the cytosolic tail was able to allow high levels of EPO sialylation. Substitution of the Gly residues with Ile in th...Continue Reading

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Citations

Mar 20, 2013·Molecular Aspects of Medicine·Zhiwei Song
Sep 10, 2013·Chembiochem : a European Journal of Chemical Biology·Andrea MaggioniJoe Tiralongo
Jun 11, 2011·Chembiochem : a European Journal of Chemical Biology·Ngalle Eric MbuaGeert-Jan Boons
Jun 4, 2010·Biotechnology and Bioengineering·Danny Chee Furng WongMiranda Gek Sim Yap
Jul 9, 2016·Glycoconjugate Journal·Roberta Salinas-MarínIván Martínez-Duncker
Sep 20, 2016·Biotechnology and Bioengineering·Joaquina X MascarenhasKevin J Kayser
Apr 28, 2018·International Journal of Molecular Sciences·Sandra BrasilVanessa Dos Reis Ferreira
Aug 30, 2019·Computational and Structural Biotechnology Journal·Barbara HadleyJoe Tiralongo
Jan 6, 2021·International Journal of Molecular Sciences·Bożena SzulcDorota Maszczak-Seneczko
Jul 14, 2011·Chemical Reviews·Ryan M SchmaltzChi-Huey Wong
Aug 14, 2021·The Journal of Biological Chemistry·Danyang Li, Somshuvra Mukhopadhyay

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