An estimation of the size of the water cluster present at the cleavage site of the water splitting enzyme

In time‐dependent measurements of oxygen evolution in tobacco thylakoid membranes we varied the fraction of H 2 18 O and the temperature and measured water splitting as 18 O 2 , 16 O 18 O, and 16 O 2 by mass spectrometry. We show that the approach to the equilibrium of the system after H 2 18 O addition can be very well understood in terms of the diffusion of water molecules. The equilibrium states of 16 O 2 , 16 O 18 O, and 18 O 2 evolution differ from the theoretical binomial distributions, which are expected under the prerequisite of ideal mixing of the water molecules and that of the chemical equivalence of H 2 18 O and H 2 16 O for an infinite cluster. The presence of this deviation means that there is a typical size of water clusters having access to cleavage by the water splitting enzyme. We estimated that this cluster contains about 12±2 water molecules.