'Molten-globule' state accumulates in carbonic anhydrase folding |
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Authors: | D A Dolgikh A P Kolomiets I A Bolotina O B Ptitsyn |
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Institution: | 1. Institute of Protein Research, USSR Academy of Sciences, 142292 Pushchino, Moscow Region, USSR;2. Institute of Molecular Biology, USSR Academy of Sciences, 117312 Moscow, USSR |
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Abstract: | Binding characteristics of 3H]BAY K 8644, a new class of pharmacologically potent compounds, the calcium channel activating dihydropyridines (DHP), were demonstrated in cultured myocardial cells. 3H]BAY K 8644 exhibited reversible and saturable binding to myocytes, and specific binding was Ca2+-dependent. The equilibrium dissociation constant, Kd, was 35.2 nM, and maximal binding capacity, Bmax, was 1.07 pmol/mg protein. Binding of the 3H-ligand was highly specific for various potently displacing DHP derivatives (either the calcium channel activating BAY K 8644, or the Ca2+ entry blockers of the nifedipine type) with inhibition constants (Ki values) in the nanomolar range. BAY K 8644, on the other hand, showed very low affinity to other receptors tested in brain and heart membranes. Displacement potency of BAY K 8644 correlated well with data of the functional pharmacology; e.g., the enhanced myocardial contractility. Results from competition studies using 3H]BAY K 8644 and 3H]nimodipine support the conclusion that both the channel activating and inhibiting DHP structures interact with the same specific receptor site that might be associated with the putative Ca2+-channel. |
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Keywords: | Carbonic anhydrase B Protein folding Molten-globule state of protein molecules Folding intermediate CA B carbonic anhydrase B α-LA α-lactalbumin Gu-HCl guanidine hydrochloride CD circular dichroism UV ultraviolet states of CA B — N (native) I (intermediate) and U (unfolded) |
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