The S-3 state of the Mn4CaO5-cluster in photosystem II was investigated by DFT calculations and compared with EXAFS data. Considering previously proposed mechanism; a water molecule is inserted into an open coordination site of Mn upon S-2 to S-3 transition that becomes a substrate water, we examined if the water insertion is essential for the S-3 formation, or if one cannot eliminate other possible routes that do not require a water insertion at the S-3 stage. The novel S-3 state structure consisting of only short 2.7-2.8 angstrom Mn Mn distances was discussed. (C) 2016 The Authors. Published by Elsevier B.V. This is an open access article under the CC BY-NC-ND license (http://creativecommons.org/licenses/by-nc-nd/4.0/).