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dc.contributor.authorHe, Xu
dc.contributor.authorPierce, Owen
dc.contributor.authorHaselhorst, Thomas
dc.contributor.authorKolarich, Daniel
dc.contributor.authorPacker, Nicolle H.
dc.contributor.authorGloster, Tracey
dc.contributor.authorVocadlo, David J.
dc.contributor.authorQian, Yi
dc.contributor.authorBrooks, Doug
dc.contributor.authorKermode, Allison R.
dc.date.accessioned2013-08-05T14:01:07Z
dc.date.available2013-08-05T14:01:07Z
dc.date.issued2013-12
dc.identifier.citationHe , X , Pierce , O , Haselhorst , T , Kolarich , D , Packer , N H , Gloster , T , Vocadlo , D J , Qian , Y , Brooks , D & Kermode , A R 2013 , ' Characterization and downstream mannose phosphorylation of human recombinant α-L-iduronidase produced in Arabidopsis complex glycan-deficient (cgl) seeds ' , Plant Biotechnology Journal , vol. 11 , no. 9 , pp. 1034–1043 . https://doi.org/10.1111/pbi.12096en
dc.identifier.issn1467-7644
dc.identifier.otherPURE: 62940362
dc.identifier.otherPURE UUID: ae658d84-9878-4323-a023-d7c7e25a5ba7
dc.identifier.otherScopus: 84888136369
dc.identifier.urihttp://hdl.handle.net/10023/3910
dc.descriptionThis work was supported by a Wellcome Trust award to TMG.en
dc.description.abstractMucopolysaccharidosis (MPS) I is a lysosomal storage disease caused by a deficiency of α-L-iduronidase (IDUA) (EC 3.2.1.76); enzyme replacement therapy is the conventional treatment for this genetic disease. Arabidopsis cgl mutants are characterized by a deficiency of the activity of N-acetylglucosaminyl transferase I (EC 2.4.1.101), the first enzyme in the pathway of hybrid and complex N-glycan biosynthesis. To develop a seed-based platform for the production of recombinant IDUA for potential treatment of MPS I, cgl mutant seeds were generated to express human IDUA at high yields and to avoid maturation of the N-linked glycans on the recombinant human enzyme. Enzyme kinetic data showed that cgl-IDUA has similar enzymatic properties to the commercial recombinant IDUA derived from cultured Chinese hamster ovary (CHO) cells (AldurazymeTM). The N-glycan profile showed that cgl-derived IDUA contained predominantly high-mannose-type N-glycans (94.5%), and the residual complex/hybrid N-glycan-containing enzyme was efficiently removed by an additional affinity chromatography step. Furthermore, purified cgl-IDUA was amenable to sequential in vitro processing by soluble recombinant forms of the two enzymes that mediate the addition of the mannose-6-phosphate (M6P) tag in mammalian cells—UDP-GlcNAc:lysosomal enzyme N−acetylglucosamine (GlcNAc)−1−phosphotransferase—and GlcNAc−1−phosphodiester α−N−acetylglucosaminidase (the ‘uncovering enzyme’). Arabidopsis seeds provide an alternative system for producing recombinant lysosomal enzymes for enzyme replacement therapy; the purified enzymes can be subjected to downstream processing to create the M6P, a recognition marker essential for efficient receptor-mediated uptake into lysosomes of human cells.
dc.language.isoeng
dc.relation.ispartofPlant Biotechnology Journalen
dc.rights© 2013 The Authors. Plant Biotechnology Journal published by Society for Experimental Biology, Association of Applied Biologists and John Wiley & Sons Ltd This is an open access article under the terms of the Creative Commons Attribution License, which permits use, distribution and reproduction in any medium, provided the original work is properly cited.en
dc.subjectArabidopsis cgl mutanten
dc.subjectMucopolysaccharidosis Ien
dc.subjectHuman α-L-iduronidaseen
dc.subjectMannose-6-phosphate recognition markeren
dc.subjectN-glycosylationen
dc.subjectQK Botanyen
dc.subjectQH426 Geneticsen
dc.subject.lccQKen
dc.subject.lccQH426en
dc.titleCharacterization and downstream mannose phosphorylation of human recombinant α-L-iduronidase produced in Arabidopsis complex glycan-deficient (cgl) seedsen
dc.typeJournal articleen
dc.description.versionPublisher PDFen
dc.contributor.institutionUniversity of St Andrews.School of Biologyen
dc.contributor.institutionUniversity of St Andrews.Biomedical Sciences Research Complexen
dc.identifier.doihttps://doi.org/10.1111/pbi.12096
dc.description.statusPeer revieweden


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