Genetic disruption of multiple α1,2-mannosidases generates mammalian cells producing recombinant proteins with high-mannose-type N -glycans

The Journal of Biological Chemistry
Ze-Cheng JinMorihisa Fujita

Abstract

Recombinant therapeutic proteins are becoming very important pharmaceutical agents for treating intractable diseases. Most biopharmaceutical proteins are produced in mammalian cells because this ensures correct folding and glycosylation for protein stability and function. However, protein production in mammalian cells has several drawbacks, including heterogeneity of glycans attached to the produced protein. In this study, we established cell lines with high-mannose-type N-linked, low-complexity glycans. We first knocked out two genes encoding Golgi mannosidases (MAN1A1 and MAN1A2) in HEK293 cells. Single knockout (KO) cells did not exhibit changes in N-glycan structures, whereas double KO cells displayed increased high-mannose-type and decreased complex-type glycans. In our effort to eliminate the remaining complex-type glycans, we found that knocking out a gene encoding the endoplasmic reticulum mannosidase I (MAN1B1) in the double KO cells reduced most of the complex-type glycans. In triple KO (MAN1A1, MAN1A2, and MAN1B1) cells, Man9GlcNAc2 and Man8GlcNAc2 were the major N-glycan structures. Therefore, we expressed two lysosomal enzymes, α-galactosidase-A and lysosomal acid lipase, in the triple KO cells and found that the g...Continue Reading

References

Jul 1, 1977·The Journal of General Virology·F L GrahamR Nairn
Oct 15, 1993·European Journal of Biochemistry·E BauseB Schmidt
May 1, 1993·The Journal of Clinical Investigation·Y Sato, E Beutler
Nov 3, 2004·Archives of Biochemistry and Biophysics·Kiyotaka FujitaKenji Yamamoto
May 3, 2005·Journal of Pharmacological and Toxicological Methods·Philip Thomas, Trevor G Smart
Jan 25, 2006·The Journal of Biological Chemistry·Kazuyoshi HiraoNobuko Hosokawa
Nov 23, 2006·Methods in Enzymology·Steven W Mast, Kelley W Moremen
Jul 29, 2008·Journal of Pharmaceutical Sciences·Ricardo J Solá, Kai Griebenow
Dec 3, 2008·Biochimica Et Biophysica Acta·Thomas Braulke, Juan S Bonifacino
Jan 13, 2010·Glycobiology·Nobuko HosokawaAnnette Herscovics
Jan 13, 2010·MAbs·Naoko Yamane-Ohnuki, Mitsuo Satoh
May 15, 2010·Analytical Chemistry·Martin StrohalmVladimír Havlícek
Apr 21, 2011·Human Genomics·Mirella Filocamo, Amelia Morrone
Sep 20, 2011·Clinical Investigation·Thomas A Burrow, Gregory A Grabowski
Sep 29, 2011·Current Opinion in Pediatrics·Robin H Lachmann
Dec 14, 2011·Applied Microbiology and Biotechnology·Jee Yon KimGyun Min Lee
Jan 5, 2013·Science·Le CongFeng Zhang
Jan 5, 2013·Science·Prashant MaliGeorge M Church
Dec 19, 2013·PLoS Genetics·Daisy RymenGert Matthijs
Feb 1, 2014·Nature Methods·Florian HeigwerMichael Boutros
Mar 15, 2014·The Journal of Biological Chemistry·Michael J IannottiRichard N Sifers
Apr 23, 2014·Nature Biotechnology·Leander MeurisNico Callewaert
Aug 6, 2014·The Journal of Cell Biology·Satoshi NinagawaKazutoshi Mori

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Citations

Oct 12, 2018·FASEB Journal : Official Publication of the Federation of American Societies for Experimental Biology·Guowei ZhangJianguo Gu
May 19, 2019·Journal of Biochemistry·Wei-Wei RenMorihisa Fujita
May 19, 2020·Cell Adhesion & Migration·Miguel Gonzalez-AndradesPablo Argüeso
Oct 14, 2020·Proceedings of the National Academy of Sciences of the United States of America·Daniel W HeindelLara K Mahal
Mar 18, 2021·Developmental Cell·Yi-Fan HuangMorihisa Fujita
Apr 21, 2021·Journal of Biochemistry·Ji-Xiong LengMorihisa Fujita

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