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- Title
A bacterial glycosidase enables mannose-6-phosphate modification and improved cellular uptake of yeast-produced recombinant human lysosomal enzymes.
- Authors
Tiels, Petra; Baranova, Ekaterina; Piens, Kathleen; De Visscher, Charlotte; Pynaert, Gwenda; Nerinckx, Wim; Stout, Jan; Fudalej, Franck; Hulpiau, Paco; Tännler, Simon; Geysens, Steven; Van Hecke, Annelies; Valevska, Albena; Vervecken, Wouter; Remaut, Han; Callewaert, Nico
- Abstract
Lysosomal storage diseases are treated with human lysosomal enzymes produced in mammalian cells. Such enzyme therapeutics contain relatively low levels of mannose-6-phosphate, which is required to target them to the lysosomes of patient cells. Here we describe a method for increasing mannose-6-phosphate modification of lysosomal enzymes produced in yeast. We identified a glycosidase from C. cellulans that 'uncaps' N-glycans modified by yeast-type mannose-Pi-6-mannose to generate mammalian-type N-glycans with a mannose-6-phosphate substitution. Determination of the crystal structure of this glycosidase provided insight into its substrate specificity. We used this uncapping enzyme together with ?-mannosidase to produce in yeast a form of the Pompe disease enzyme ?-glucosidase rich in mannose-6-phosphate. Compared with the currently used therapeutic version, this form of ?-glucosidase was more efficiently taken up by fibroblasts from Pompe disease patients, and it more effectively reduced cardiac muscular glycogen storage in a mouse model of the disease.
- Subjects
GLYCOSIDASES; MANNOSE 6-phosphate; LYSOSOMAL storage diseases; THERAPEUTICS; LYSOSOMES; MAMMALIAN cell cycle; YEAST; GLYCANS; CRYSTAL structure; GLYCOGEN storage disease; LABORATORY mice
- Publication
Nature Biotechnology, 2012, Vol 30, Issue 12, p1225
- ISSN
1087-0156
- Publication type
Article
- DOI
10.1038/nbt.2427