<title>Nonexponential decay of room-temperature phosphorescence: evidence for several slowly interconverting or static protein conformers</title>

Abstract

The phosphorescence decays of horse liver alcohol dehydrogenase (LADH) Trp314, E. coli alkaline phosphatase (AP) Trp109, and B. stearothermophilus phosphofructokinase (PFK) Trp179 are decidedly nonexponential at room temperature. When the data is analyzed using the maximum entropy method (MEM) the AP phosphorescence decay is dominated by a single gaussian distribution while for LADH and PFK the data reveals at least two amplitude packets. The MEM lifetime-normalized widths for these proteins are significantly larger than obtained for the model monoexponential chromophore terbium suggesting that the complex kinetics is intrinsic to the protein. Since the phosphorescence lifetime of a tryptophan residue is related to its microviscosity, the nonexponential decay behavior may imply that the phosphorescing tryptophan residue in each of these samples is best described as existing in at least two states of different local rigidity which interconvert more slowly than the time scale of the phosphorescence decay (0.1 to 1.0 sec). The existence of multiple, long-lived, conformers is further supported by the observation that the phosphorescence lifetime in the LADH sample is excitation wavelength dependent.