Interaction of rabbit muscle pyruvate kinase (ATP:pyruvate 2-O-phosphotransferase, EC 2.7.1.40) with inhibitory and activatory effectors was analyzed in terms of the Yagi-Ozawa equation. Straight-line plots obtained for L-phenylalanine + L-alanine and L-phenylalanine + L-phenylalanyl methyl ester (PheAlaOMe) indicated competition for common sites. Parabolic plots for 2-phosphoglycerate and L-phenylalanine or PheAlaOMe suggested lack of competition between these compounds. Biphasic concave plots were obtained for 2-phosphoglycerate and L-1-amino-2-phenylethyl phosphonic acid (PnPhe), and mixtures of inhibitory (L-phenylalanine, PheAlaOMe) and activatory (PnPhe, L-alanine and L-1-aminoethyl phosphonic acid (PnAla)) amino acid derivatives. The latter non-classical Yagi-Ozawa plots can be interpreted in terms of the existence of distinct regulatory sites for inhibitory and activatory ligands.