X-ray absorption spectroscopy (XAS) has been used to characterize the structural consequences of Ca2+ replacement in the reaction center complex of the photosynthetic oxygen-evolving complex (OEC). EPR and activity measurements demonstrate that, in the absence of the 17 and 23 kDa extrinsic polypeptides, it is nor necessary to use either low pH or Ca chelators to effect complete replacement of the active site Ca2+ by Sr2+, DY3+, or La3+. The extended X-ray absorption fine structure (EXAFS) spectra for the OEC show evidence for a Mn ... M interaction at ca. 3.3 Angstrom that could arise either from Mn ... Mn scattering within the Mn cluster or Mn ... Ca scattering between the Mn cluster and the inorganic Ca2+ cofactor. There is no significant change in the either the amplitude or the phase of this feature when Ca2+ is replaced by Sr2+ or DY3+, thus demonstrating that there is no EXAFS-detectable Mn ... Ca contribution at ca. 3.3 Angstrom in these samples. The only significant consequence of Ca2+ replacement is a small change in the ca. 2.7 Angstrom Mn ... Mn distance. The average Mn ... Mn distance decreases 0.014 Angstrom when Ca2+ is replaced by Sr2+ and increases 0.012 Angstrom when Ca2+ is replaced by Dy3+. A structural model which can account both for the variation in Mn ... Mn distance and for the known properties of Ca2+-substituted samples is one in which there is a hydrogen bond between a Ca2+-bound water and a Mn-2(mu-O)(2) unit. This scheme suggests that an important role for the Ca2+ may be to modulate the protonation stare, and thus the redox potential, of the Mn cluster.