title: Functional Analysis of the Ser149/Thr149 Variants of Human Aspartylglucosaminidase and Optimization of the Coding Sequence for Protein Production.
abstract: Aspartylglucosaminidase (AGA) is a lysosomal hydrolase that participates in the breakdown of glycoproteins. Defects in the AGA gene result in a lysosomal storage disorder, aspartylglucosaminuria (AGU), that manifests mainly as progressive mental retardation. A number of AGU missense mutations have been identified that result in reduced AGA activity. Human variants that contain either Ser or Thr in position 149 have been described, but it is unknown if this affects AGA processing or activity. Here, we have directly compared the Ser149/Thr149 variants of AGA and show that they do not differ in terms of relative specific activity or processing. Therefore, Thr149 AGA, which is the rare variant, can be considered as a neutral or benign variant. Furthermore, we have here produced codon-optimized versions of these two variants and show that they are expressed at significantly higher levels than AGA with the natural codon-usage. Since optimal AGA expression is of vital importance for both gene therapy and enzyme replacement, our data suggest that use of codon-optimized AGA may be beneficial for these therapy options.
authors.affiliation: Institute of Biochemistry, Medical Faculty, University of Giessen, Friedrichstrasse 24, 35392 Giessen, Germany. Antje.Banning@biochemie.med.uni-giessen.de.
authors.forename: Antje
authors.initials: A
authors.lastname: Banning
authors.affiliation: Institute of Biochemistry, Medical Faculty, University of Giessen, Friedrichstrasse 24, 35392 Giessen, Germany. jan.friedrich.koenig@googlemail.com.
authors.forename: Jan F
authors.initials: JF
authors.lastname: König
authors.affiliation: Gene Therapy Center and Department of Ophthalmology, University of North Carolina, Chapel Hill, NC 27302, USA. Steven_Gray@med.unc.edu.
authors.forename: Steven J
authors.initials: SJ
authors.lastname: Gray
authors.affiliation: Institute of Biochemistry, Medical Faculty, University of Giessen, Friedrichstrasse 24, 35392 Giessen, Germany. Ritva.Tikkanen@biochemie.med.uni-giessen.de.
authors.forename: Ritva
authors.initials: R
authors.lastname: Tikkanen
mesh_terms: D001227:Aspartylglucosylaminase / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D002478:Cells, Cultured; D005347:Fibroblasts / Q000166:cytology / Q000378:metabolism / Q000473:pathology; D005787:Gene Frequency; D005838:Genotype; D057809:HEK293 Cells; D006367:HeLa Cells; D006801:Humans; D016464:Lysosomal Storage Diseases / Q000201:enzymology / Q000235:genetics / Q000473:pathology; D008247:Lysosomes / Q000737:chemistry / Q000378:metabolism; D010957:Plasmids / Q000235:genetics / Q000378:metabolism; D020641:Polymorphism, Single Nucleotide; D021122:Protein Subunits / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D014162:Transfection
chemical_list: D021122:Protein Subunits; D001227:Aspartylglucosylaminase
keywords: aspartylglucosaminidase; aspartylglucosaminuria; gene therapy; lysosomal storage disorder; lysosomes; single nucleotide polymorphism