Investigation of substrate water interactions at the high-affinity Mn site in the photosystem II oxygen-evolving complex

Abstract

18O isotope exchange measurements of photosystem II (PSII ) in thylakoids from wild-type and mutant Synechocystis have been performed to investigate binding of substrate water to the highaffinity Mn 4 site in the oxygen-evolving complex (OEC). The mutants investigated were D1-D170H, a mutation of a direct ligand to the Mn 4 ion, and D1-D61N, a mutation in the second coordination sphere. The substrate water 18 O exchange rates for D61N were found to be 0.16G0.02 s K1 and 3.03G0.32 s K1 for the slow and fast phases of exchange, respectively, compared with 0.47G0.04 s K1 and 19.7G1.3 s K1 for the wild-type. The D1-D170H rates were found to be 0.70G0.16 s K1 and 24.4G4.6 s K1 and thus are almost within the error limits for the wild-type rates. The results from the D1-D170H mutant indicate that the high-affinity Mn 4 site does not directly bind to the substrate water molecule in slow exchange, but the binding of non-substrate water to this Mn ion cannot be excluded. The results from the D61N mutation show an interaction with both substrate water molecules, which could be an indication that D61 is involved in a hydrogen bonding network with the substrate water. Our results provide limitations as to where the two substrate water molecules bind in the OEC of PSII.

Oxygen-evolving complex of Photosystem II: an analysis of second-shell residues and hydrogen-bonding networks

“…The D1-D61N mutation slows down both W s and W f exchange rates by 3 and 6.5 times, respectively [20]. This is especially surprising because mutations of some of the direct ligands to the OEC, such as D1-D170 and D1-E189, have less …”

Uncovering channels in photosystem II by computer modelling: current progress, future prospects, and lessons from analogous systems

“…Additionally, the residues D1-D61 and CP43-R357, which have been proposed to be important in the deprotonation steps during the S-cycle (Dau and Haumann 2008;Knoepfle et al 1999;McEvoy and Brudvig 2004;McEvoy et al 2005;Sproviero et al 2006Sproviero et al , 2008a form parts of these channels. More recently, these residues have also been shown by mutagenesis studies to be very important to optimal S-cycling and O 2 evolution, even without being ligands to the CaMn 4 cluster (Debus 2005;Hwang et al 2007;Singh et al 2008 channels, the assignment of the large channel system for O 2 exit was preferred, as its volume and numerous exits may facilitate rapid removal of O 2 compared to the much narrower back channel. Also, it was found by comparison with cryogenic electron microscopy of PSII supercomplexes that the large channel system would direct O 2 away from the associated chlorophyll-rich subunits and light harvesting complexes.…”

Substrate water binding and oxidation in photosystem II

“…This work has indicated that H-bonding is critical in the binding process (Noguchi andSugiura 2000, 2002b). Interestingly water exchange measurements conducted in the S state of a D1:D170H mutant show no significant difference in the rates of exchange of either fast or slow exchanging water sites between the mutant and wild-type PSII (Singh et al 2008). Surprisingly, this indicates that D170 mostly likely does not ligate a Mn that binds a substrate water molecule as D170 is thought to ligate the dangler Mn4 and is widely considered to be a substrate water binding site.…”