Electron transport in Photosystem II of higher plants is particularly susceptible to inhibition at the site where electrons are transferred between the plastoquinone electron acceptors Q_A and Q_B. Substituted benzoquinones, naphthoquinones, anthraquinones, coumarins, and pyrones, as structural analogs of Q_B, compete with Q_B for a common binding domain and inhibit electron transfer from Q_A. We have used measurements of chlorophyll fluorescence intensity as well as CNDO molecular orbital calculations to (1) characterize the inhibitor strength; (2) correlate the chemical structure of the inhibitors to the extent of inhibitory activity; and (3) predict the binding site of the inhibitors on the Q_B-binding protein. Stern-Volmer analyses of the chlorophyll fluroscence data provide two parameters for assessing inhibitor strength. These parameters suggest that the variable inhibitory activities observed for the Q_B analogs reflect both the hydrophobicity and the electron-releasing character of the substituents. CNDO calculations reveal that inhibitors with the same π charge distribution on the atoms adjacent to the carbonyl carbons compete for a common binding domain. These studies permit the prediction of both the substituent identity and the substitution pattern in quinones, coumarins, and pyrones that lead to maximal inhibition of photosynthetic activity.