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Kramer, Werner; Müller, Günter; Girbig, Frank; Gutjahr, Ulrike; Kowalewski, Simone; Hartz, Detlev und Summ, Hans-Dieter (11. Mai 1994): Differential interaction of glimepiride and glibenclamide with the β-cell sulfonylurea receptor II. Photoaffinity labeling of a 65 kDa protein by [3H]glimepiride. In: Biochimica et Biophysica Acta (BBA) - Biomembranes, Vol. 1191, Nr. 2: S. 278-290




Glimepiride is a novel sulfonylurea for the treatment of type II-diabetic patients exhibiting different receptor binding kinetics to β-cell membranes with 8–9-fold higher koff rate and 2.5–3-fold higher kon rate compared to glibenclamide (see accompanying paper (Müller, G. et al. (1994) Biochim. Biophys. Acta 1191, 267–277)). To elucidate the molecular basis for this differential behaviour of glimepiride and glibenclamide, direct photoaffinity labeling studies using β-cell tumor membranes were performed. [3H]Glimepiride was specifically incorporated into a membrane polypeptide of Mr = 65000 under conditions, which led to predominant labeling of a 140 kDa protein by [3H]glibenclamide (Kramer, W. et al. (1988) FEBS Lett. 229, 355–359). Labeling of the 140 kDa protein by [3H]glibenclamide was inhibited by unlabeled glimepiride and, vice versa, glibenclamide inhibited labeling of the 65 kDa protein by [3H]glimepiride. The 65 kDa protein was also specifically photolabeled by the sulfonylurea [125I]35623, whereas an 4-azidobenzoyl derivative of glibenclamide, N3-[3H]33055, exclusively labeled a 33 kDa protein. Competitive Scatchard analysis of [3H]glimepiride-binding and [3H]glibenclamide-binding to RINm5F cell membranes using glibenclamide and glimepiride, respectively, as heterologous displacing compounds yielded non-linear plots. These findings may be explained by cooperative interactions between the 140 and 65 kDa sulfonylurea-binding proteins. The possibility that sulfonylureas of different structure have different access to the 140 and 65 kDa receptor proteins due to the β-cell membrane barrier was investigated by photoaffinity labeling of solubilized β-cell membrane proteins. Interestingly, solubilization of β-cell tumor membranes led to a shift of specific [3H]glibenclamide binding from the 140 kDa to the 65 kDa binding protein, exclusively, and to an increased labeling of the 65 kDa protein by [3H]glimepiride. The labeling of a unique protein is in agreement with similar Kd values measured for both sulfonylurcas upon solubilization of β-cell tumor and RINm5F cell membranes (see accompanying paper). Furthermore, competitive Scatchard plots of [3H]glimepiride binding to solubilized RINm5F cell membrane proteins in the presence of glibenclamide and vice versa approximate linearity suggesting loss of cooperativity between the 140 kDa glibenclamide-binding and 65 kDa glimepiride-binding proteins upon solubilization. The physiological significance of the differential interaction of glimepiride and glibenclamide with different binding proteins was also substantiated by photoaffinity labeling of RINm5F cells leading to labeling of a 140 kDa protein by [3H]glibenclamide and of a 65 kDa protein by [3H]glimepiride. In conclusion, this report presents the first evidence that different sulfonylurea drugs bind to different components of the sulfonylurea receptor complex which are characterized by different accessibility for the drugs.