Article ID | Journal | Published Year | Pages | File Type |
---|---|---|---|---|
10956108 | Molecular and Cellular Endocrinology | 2014 | 11 Pages |
Abstract
GCK-MODY, dominantly inherited mild hyperglycemia, is associated with more than 600 mutations in the glucokinase gene. Different molecular mechanisms have been shown to explain GCK-MODY. Here, we report a Pakistani family harboring the glucokinase mutation c.823C > T (p.R275C). The recombinant and in cellulo expressed mutant pancreatic enzyme revealed slightly increased enzyme activity (kcat) and normal affinity for α-D-glucose, and resistance to limited proteolysis by trypsin comparable with wild-type. When stably expressed in HEK293 cells and MIN6 β-cells (at different levels), the mutant protein appeared misfolded and unstable with a propensity to form dimers and aggregates. Its degradation rate was increased, involving the lysosomal and proteasomal quality control systems. On mutation, a hydrogen bond between the R275 side-chain and the carbonyl oxygen of D267 is broken, destabilizing the F260-L271 loop structure and the protein. This promotes the formation of dimers/aggregates and suggests that an increased cellular degradation is the molecular mechanism by which R275C causes GCK-MODY.
Keywords
GCKpost nuclear supernatantα-d-glucoseRRLGKRPhGKDulbecco’s modified eagles mediumIFGHEKDMEMPDBGlcBCAImpaired fasting glucosebicinchoninic acidAggregationMinSelf-associationMaturity-onset diabetes of the youngrabbit reticulocyte lysateProtein misfoldingCatalytic activityMODYProtein Data BankGlucokinase regulatory proteinPNShuman embryonic kidney
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Authors
Maria Negahdar, Ingvild Aukrust, Janne Molnes, Marie H. Solheim, Bente B. Johansson, Jørn V. Sagen, Knut Dahl-Jørgensen, Rohit N. Kulkarni, Oddmund Søvik, Torgeir Flatmark, Pål R. Njølstad, Lise Bjørkhaug,