Tryptophan Dynamics and Cooperativity in ATCase from E. coli
The temperature and viscosity dependence of the rotations of the intrinsic tryptophan moieties in ATCase from E. coli was studied as a function of temperature. The two tryptophan residues were differentiated in the decay curve analysis, as a short lifetime (1-3 ns) and a long lifetime (4-6 ns) component were found. The two protein slopes observed in the Y-plots were interpreted as corresponding to the two tryptophan residues which were differentiated by their surrounding free space. Using the fractional intensity data obtained from fluorescence intensity decay measurements the observed slopes were decomposed into the protein slopes for each tryptophan residue. In ATCase alone, the two were similarly coupled to the protein matrix. However, upon binding of the substrate analogue, PALA, one tryptophan was decoupled from the protein, whereas the other appeared to be more restrained. The fixation of the nucleotide effector molecules elicited contrary results, despite the fact that they bind to the same site. ATP increases the free space around one tryptophan and slightly decoupled both their motions, whereas CTP binding gave similar results as those obtained in presence of PALA. These studies have shown that the fixation of small molecules at a large distance from a given amino-acid residue can have important and specific effects upon its dynamic properties.