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Schlagwörter:
Anions, Asparagine, Aspartic Acid, Azides, Bacteriorhodopsins, Biological Transport, Hydrogen-Ion Concentration, Mutation, Protons, Temperature, Thermodynamics
Zusammenfassung:
Addition of azide fully restored the proton pump activity of defective bacteriorhodopsin (BR) mutant protein Asp96 → Asn. The decay time of M of BR Asp96 → Asn, the longest living intermediate, was decreased from 500 ms at pH 7.0 to approximately 1 ms under conditions of saturating azide concentrations. This decay was faster than the decay of M in the wild-type, where no such azide effect was detectable. Stationary photocurrents, measured with purple membranes immobilized and oriented in a polyacrylamide gel, increased upon addition of azide up to the level of the wild-type. Different small anions of weak acids restored the pump activity with decreasing affinity in the order: cyanate greater than azide greater than nitrite greater than formiate greater than acetate. The activation energy of the M decay in the mutant was higher in the presence (48 kJ/mol) than in the absence (27 kJ/mol) of 100 mM azide even though the absolute rate was dramatically increased by azide. This effect of azide is due to the substitution of a carboxamido group for a carboxylic group at position 96 which removes the internal proton donor and causes an increase in the entropy change of activation for proton transfer which is reversed by azide.
