title: Molecular pathogenesis of a new glycogenosis caused by a glycogenin-1 mutation.
abstract: Glycogenin-1 initiates the glycogen synthesis in skeletal muscle by the autocatalytic formation of a short oligosaccharide at tyrosine 195. Glycogenin-1 catalyzes both the glucose-O-tyrosine linkage and the α1,4 glucosidic bonds linking the glucose molecules in the oligosaccharide. We recently described a patient with glycogen depletion in skeletal muscle as a result of a non-functional glycogenin-1. The patient carried a Thr83Met substitution in glycogenin-1. In this study we have investigated the importance of threonine 83 for the catalytic activity of glycogenin-1. Non-glucosylated glycogenin-1 constructs, with various amino acid substitutions in position 83 and 195, were expressed in a cell-free expression system and autoglucosylated in vitro. The autoglucosylation was analyzed by gel-shift on western blot, incorporation of radiolabeled UDP-(14)C-glucose and nano-liquid chromatography with tandem mass spectrometry (LC/MS/MS). We demonstrate that glycogenin-1 with the Thr83Met substitution is unable to form the glucose-O-tyrosine linkage at tyrosine 195 unless co-expressed with the catalytically active Tyr195Phe glycogenin-1. Our results explain the glycogen depletion in the patient expressing only Thr83Met glycogenin-1 and why heterozygous carriers without clinical symptoms show a small proportion of unglucosylated glycogenin-1.
authors.affiliation: Department of Pathology, Institute of Biomedicine, University of Gothenburg, Gothenburg, Sweden. johanna.nilsson@gu.se
authors.forename: Johanna
authors.initials: J
authors.lastname: Nilsson
authors.forename: Adnan
authors.initials: A
authors.lastname: Halim
authors.forename: Ali-Reza
authors.initials: AR
authors.lastname: Moslemi
authors.forename: Anders
authors.initials: A
authors.lastname: Pedersen
authors.forename: Jonas
authors.initials: J
authors.lastname: Nilsson
authors.forename: Göran
authors.initials: G
authors.lastname: Larson
authors.forename: Anders
authors.initials: A
authors.lastname: Oldfors
mesh_terms: D000595:Amino Acid Sequence; D001483:Base Sequence; D055162:Biocatalysis; D002474:Cell-Free System; D002850:Chromatography, Gel; D002853:Chromatography, Liquid; D017931:DNA Primers; D005947:Glucose / Q000378:metabolism; D005964:Glucosyltransferases / Q000737:chemistry / Q000235:genetics* / Q000302:isolation & purification; D006003:Glycogen / Q000096:biosynthesis*; D006023:Glycoproteins / Q000737:chemistry / Q000235:genetics* / Q000302:isolation & purification; D006801:Humans; D008969:Molecular Sequence Data; D009154:Mutation; D053719:Tandem Mass Spectrometry
chemical_list: D017931:DNA Primers; D006023:Glycoproteins; C051887:glycogenin; D006003:Glycogen; D005964:Glucosyltransferases; D005947:Glucose