Revised Manuscript Received October 30, 1991 abstract: Eleven site-directed mutations were constructed at aspartate 170 of the Dl polypeptide of the photosystem II (PSII) reaction center of the cyanobacterium Synechocystis sp. PCC 6803. The light-saturated rates of 02 evolution (Vq2) measured in whole cells range from close to that of wild-type for Aspl70Glu to zero for Aspl70Ser and Ala. Those mutant strains that are best able to evolve 02 are also those that show the lowest Km in PSII core complexes for the oxidation of Mn2+ by oxidized Tyrl61, the normal oxidant of the Mn cluster responsible for 02 evolution. To a first approximation, the lower the pXa of the residue at position 170, the higher the V0l and the lower the Km. Dl-Aspl70 appears to participate in the early steps associated with the assembly of the Mn cluster. It is also the first reported example of an amino acid residue critical to the function and assembly of the oxygen-evolving complex. e reaction center of photosystem II (PSII) 1 is one of the two natural photovoltaic cells that driveelectron transfer in the electron transport chain of oxygenic photosynthesis [for a recent review, see Hansson and Wydrzynski (1990)]. Photoexcitation of the primary electron donor, P680 (a chlo-rophyll a or pair of chlorophylls a), results in an electron transfer to the primary electron acceptor, a pheophytin a. The electron is then transferred to the primary plastoquinone electron acceptor, QA, and subsequently to the secondary plastoquinone electron acceptor, QB. These reactions bear a strong kinetic and spatial resemblence to those that occur on homologous components in the reaction centers of the purple non-sulfur photosynthetic bacteria (Michel & Deisenhofer, 1989; Komiya et al., 1988). The crystallographic structure of reaction centers from two such sources has been recently solved toatomic resolution (Michel & Deisenhofer, 1989; Yeates et al., 1988; Allen et al., 1988; El-Kabbani et al., 1991). The electron donor side of the PSII reaction center is, however, quite different from that of the photosynthetic bacteria. Much higher oxidizing potentials (£ m~ 1.2 V) are generated by the oxidized form of the primary electron donor, P680, to allow the oxidation of a redox active tyrosine, Yz, which in turn oxidizes