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Título

Effects of novel maturity-onset diabetes of the young (MODY)-associated mutations on glucokinase activity and protein stability

AutorGalán, María; Vincent, Olivier ; Roncero, Isabel; Azriel, Sharona; Boix-Pallares, Pedro; Delgado-Álvarez, Elías; Díaz-Cadórniga, Fernando; Blázquez, Enrique ; Navas, María-Angeles
Palabras claveDiabetes mellitus
Enzyme kinetic
GCK gene
Glucokinase
Inactivating mutation
Maturity-onset diabetes of the young (MODY)
Fecha de publicación1-ene-2006
EditorPortland Press
CitaciónBiochemical Journal 393(1): 389-396 (2006)
ResumenGlucokinase acts as the pancreatic glucose sensor and plays a critical role in the regulation of insulin secretion by the b-cell. Heterozygous mutations in the glucokinase-encoding GCK gene, which result in a reduction of the enzymatic activity, cause the monogenic form of diabetes, MODY2 (maturity-onset diabetes of the young 2). We have identified and functionally characterized missense mutations in the GCK gene in diabetic families that result in protein mutations Leu165Phe, Glu265Lys and Thr206Met. The first two are novel GCK mutations that co-segregate with the diabetes phenotype in their respective families and are not found in more than 50 healthy control individuals. In order to measure the biochemical effects of these missense mutations on glucokinase activity, we bacterially expressed and affinity-purified islet human glucokinase proteins carrying the respective mutations and fused to GST (glutathione S-transferase). Enzymatic assays on the recombinant proteins revealed that mutations Thr206Met and Leu165Phe strongly affect the kinetic parameters of glucokinase, in agreement with the localization of both residues close to the active site of the enzyme. In contrast, mutation Glu265Lys, which has a weaker effect on the kinetics of glucokinase, strongly affects the protein stability, suggesting a possible structural defect of this mutant protein. Finally, none of the mutations tested appears to affect the interaction of gluco-kinase with the glucokinase regulatory protein in the yeast two-hybrid system.
Descripción8 pages, 4 figures, 3 tables.
Versión del editorhttp://www.biochemj.org/bj/393/0389/bj3930389.htm
URIhttp://hdl.handle.net/10261/24847
ISSN0264-6021
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