title: Human hereditary glutathione synthetase deficiency: kinetic properties of mutant enzymes.
abstract: Patients with hereditary glutathione synthetase deficiency suffer from haemolytic anaemia, 5-oxoprolinuria, metabolic acidosis, recurrent bacterial infections and various degrees of central nervous system dysfunction. To investigate the molecular basis of the mutations associated with this disease, seven naturally occurring missense mutations [L188P (Leu188-->Pro), D219A, D219G, Y270C, Y270H, R283C and P314L] were expressed using a His-tagged, Escherichia coli-based expression system. Effects of the mutations on kinetic properties, including negative co-operative binding of gamma-glutamyl substrate, were evaluated. The mutation P314L did not have any major effect on these parameters and was classified as a neutral mutation. The remaining mutations decreased V(max) to 2-27% of wild-type activity. Negative co-operativity for gamma-gluABA (L-gamma-glutamyl-L-alpha-aminobutyric acid) was abolished in five mutant recombinant enzymes, whereas for one mutant enzyme, this co-operativity changed from negative to positive. The structural consequences of the mutations were interpreted on the basis of the known structure of the wild-type enzyme.
authors.affiliation: Department of Paediatrics, Karolinska Institutet, Karolinska University Hospital, 141 86 Stockholm, Sweden. runa.njalsson@mednut.ki.se
authors.forename: Runa
authors.initials: R
authors.lastname: Njålsson
authors.forename: Katarina
authors.initials: K
authors.lastname: Carlsson
authors.forename: Vikas
authors.initials: V
authors.lastname: Bhansali
authors.forename: Jia-Li
authors.initials: JL
authors.lastname: Luo
authors.forename: Lennart
authors.initials: L
authors.lastname: Nilsson
authors.forename: Rudolf
authors.initials: R
authors.lastname: Ladenstein
authors.forename: Mary
authors.initials: M
authors.lastname: Anderson
authors.forename: Agne
authors.initials: A
authors.lastname: Larsson
authors.forename: Svante
authors.initials: S
authors.lastname: Norgren
mesh_terms: D000592:Amino Acid Metabolism, Inborn Errors / Q000201:enzymology*; D005981:Glutathione Synthase / Q000737:chemistry / Q000172:deficiency* / Q000378:metabolism* / Q000502:physiology; D006801:Humans; D008958:Models, Molecular
chemical_list: D005981:Glutathione Synthase