Switch from conventional to distributed kinetics in the bacteriorhodopsin photocycle

Below 195 K, the bacteriorhodopsin photocycle could not be adequately described with exponential kinetics [Dioumaev, A. K., and Lanyi, J. K. (2007) Proc. Nad. Acad. Sci. U.S.A. 104, 9621-9626] but required distributed kinetics, previously found in hemoglobin and myoglobin at temperatures below the vitrification point of the surrounding solvent. The aim of this study is to determine which factors cause the switch from this low-temperature regime to the conventional kinetics observed at ambient temperature. The photocycle was monitored by time-resolved FTIR between 180 and 280 K, using, the D96N mutant. Depending on the temperature, decay and temporal redistribution of two or three intermediates (L, M, and N) were observed. Above similar to 245 K, an abrupt change in the kinetic behavior of the photocycle takes place. It does not affect the intermediates present but greatly accelerates their decay. Below similar to 240 K, a kinetic pattern with partial decay that cannot be explained by conventional kinetics, but suggesting distributed kinetics, was dominant, while above similar to 250 K, there were no significant deviations from exponential behavior. The similar to 245 K critical point is >= 10 K below the freezing point of interbilayer water, and we were unable to correlate it with any FTIR-detectable transition of the lipids. Therefore, we attribute the change from distributed to conventional kinetics to a thermodynamic phase transition in the protein. Most probably, it is related to the freezing and thawing of internal fluctuations of the protein, known as the dynamic phase transition, although in bacteriorhodopsin the latter is usually believed to take place at least 15 K below the observed critical temperature of similar to 245 K.