[Intrinsic luminescence of protein as a tool for studying fast structural dynamics]

The photoexcitation of Trp, Tyr or Phe chromophore in a protein is analogous to the instant (10(-15) to 10(-14) s) introduction of a local probe carrying strong electronic and protonic donor and acceptor moieties and, moreover, having a significant dipole moment (4 to 11 D). Depending on the protein structure mobility during the excited state lifetime, the intra-macromolecular complexes (the exciplexes) with polar groups can be formed and some reversible charge transfer processes, which qaench the fluorescence, can take place with greater or lesser probability. The shifts of Trp fluorescence spectra from 307 to 316 nm (due to the 1:1 exciplex formation) and to 325-330 nm (2:1 exciplex) are typical for many proteins. An additional spectral shift up to 350 nm is caused by the reorientational relaxation of a protein matrix dipoles during the nanosecond excitation lifetimes. The bathochromic fluorescence of several proteins evidences the slow (times much longer than 10 ns) reorientation mobility in these cases. The fluorescence quenching rates by proteic groups and extrinsic small quenchers is linearly related to the diffusion coefficient of a surrounding solvent. Analysis of such relationships allows, in principle, to evaluate the diffusional internal friction inside a protein and parameters of co-operativity and anisotropy of the mobility. Some sources of possible misinterpretations of Trp fluorescence depolarisation as a measure of the rotational mobility of indole.