Fluorescence Quenching by Intrinsic Quenchers

In the case in which the protein molecule contains only a single fluorophore, the equation for the fluorescence quantum yield Q will be 

It was shown(52) that the function f(T) is not described properly by the exponent but corresponds perfectly to the temperature dependence of the T/y ratio  

Since the rate constants of bimolecular diffusion-limited reactions in isotropic solution are proportional to T/ these data testify to the fact that the kt values are linearly dependent on the diffusion coefficient D in water, irrespective of whether the fluorophores are present on the surface of the macromolecule (human serum albumin, cobra neurotoxins, proteins A and B of the neurotoxic complex of venom) or are localized within the protein matrix (ribonuclease C2, azurin, L-asparaginase).1 36 1 The linear dependence of the functions l/Q and l/xF on x/t] indicates that the mobility of protein structures is correlated with the motions of solvent molecules, and this correlation results in similar mechanisms of quenching for both surface and interior sites of the macromolecule. 

The motions of chromophore groups and of their environment that lead to temperature-dependent fluorescence quenching are those on the nanosecond time scale. Slower motions cannot manifest themselves in effects on the excited-state lifetime (this corresponds to the limit of high viscosity). On the other hand, if the motions are considerably faster (on the picosecond time scale), then they should give rise to static quenching. 

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