Enzymatic characterization of four new mutations in the glucose-6 phosphatase (G6PC) gene which cause glycogen storage disease type 1a

Glycogen storage disease type la (GSD1a) is caused by mutations in the gene of glucose-6 phosphatase (G6PC), encoding the last enzyme of gluconeogenes is and glycogenolysis. To study the effect of mutations previously identifi ed, but not yet enzymatically characterized, in French GSD1a patients, we u sed an in vitro expression system of the human glucose-6 phosphatase (hGlc6 Pase) cDNA. Wild type hGlc6Pase expressed in COS-7 cells exhibited kinetic features comparable to microsomal Glc6Pase from normal human liver and kidn ey. Pour new mutations inducing aminoacid changes in the coding sequence, e .g. W77R, A124T, G184E and L211P, were inserted into the Glc6Pase cDNA by s ite-directed mutagenesis, and studied after transient expression in COS-7 c ells. All four mutations totally abolished Glc6Pase activity.