Lack of photoactivation capacity in Scenedesmus obliquus LF-1 results from loss of half the high-affinity manganese-binding site : Relationship to the unprocessed D1 protein

Abstract

The high-affinity binding site for Mn 2+ is characterized by a decrease in 1,5-diphenylcarbazide (DPC) to 2,6-dichlorophenolindophenol (DCIP) electron transport with NH 2 OH-treated spinach Photosystem II (PS II) membrane fragments when micromolar amounts of Mn 2+ are present in the assay. This site is purported to be the binding site for Mn, functional in O 2 evolution (Hsu, B.-D., Lee, J.-Y. and Pan, R.-L. (1987) Biochim. Biophys. Acta 890, 89–96). We have examined this site in PS II-enriched membranes from Scenedesmus obliquus wild-type (WT) and LF-1 mutant cells. LF-1 inserts an unprocessed D1 protein into the photosynthetic membrane, binds approx. 40% of the functional Mn as WT, and does not evolve O 2 (Metz, J.G., Pakrasi, H.B., Seibert, M. and Arntzen, C.J. (1986) FEBS Lett. 205, 269–274). The dissociation constant for added high-affinity Mn 2+ is about 0.3–0.4 μM in wheat, WT, and LF-1 PS II. However, the relative amount of available high-affinity Mn 2+ -binding site is about half as much in LF-1 PS II membranes compared to wheat, spinach, and WT PS II membranes. Despite the fact that LF-1 PS II can photoligate Mn, LF-1 cannot be photoactivateid as can NH 2 OH-treated WT PS II. LF-1 subjected to photoactivating conditions does not reach S 2 as determined by thermoluminescence. This work indicates that the Hsu et al. high-affinity Mn 2+ site is actually at least two sites, one of which is missing in LF-1, and that successful photoactivation potential requires the presence of all high-affinity Mn 2+ site. The fact that the full complement of high-affinity Mn 2+ -binding site is observed in isolated spinach PS II reaction center (D1/D2/cytochrome b -559) complex demonstrates that other PS II core proteins do not affect the high-affinity site. Histidine chemical modifier experiments show that one component of the high-affinity site is probably associated with histidine(s) and that this component is missing in LF-1. We conclude that histidine(s) on the Dl protein provides ligand(s) for part of the Mn required for O 2 -evolution function and that the balance of the Mn is bound by other amino acids on the proteins composing the PS II reaction center.