Fluorescence Lifetime of Fluorescent Proteins

Fluorescence is a photophysical phenomenon, which obeys basic physical laws. The fluorescence of the autofluorescent proteins arises on the molecular level from chromophores, which are buried in the protein matrix. The three-dimensional, well-defined architecture of the surrounding is a prerequisite for their function. Excitation of the isolated chromophores leads only to a negligible light emission at room temperature. Several processes competing with the radiative decay are responsible for the quenching. To understand how nature has learned to suppress these alternative pathways from the excited state in autofluorescent proteins, the molecular dynamics as well as the influence of several amino acids in the interior of the protein has to be analysed. We review the current status of the understanding of the non-radiative decay mechanisms for the different fluorescent protein classes, i.e., colours. Furthermore, we address what can be learned from fluorescence lifetime measurements and how they can be exploited for analytical purposes such as fluorescence lifetime imaging microscopy. Finally, we sketch the needs of increased fluorescence quantum yields and present strategies to prolong the fluorescence lifetimes.