High mobility group proteins HMG1 and -2 and histone H1 are structural components of chromatin. Previously, we reported that HMG1 interacts with H1 histone in a way that modulates the ability of H1 to condense DNA in vitro, suggesting that these proteins may act together in vivo to regulate locally the condensation state of chromatin, possibly affecting replication and/or transcription. Here we show that reduced (native) HMG1 binds to H1 cooperatively at pH 6.0 as a tetramer with a dissociation constant of 3.4 x 10(-8) M, and at pH 7.5 as a monomer with a dissociation constant less than 10(-9) M. Denaturation through oxidation of sulfhydryl groups has a strong effect on the interaction of HMG1 with H1 histone, suggesting that the reduced state of HMG1 is critical to its function. Oxidized HMG1 failed to bind H1 at pH 7.5, and its binding at pH 6 was biphasic; the first three (or two) molecules of H1 were bound with a dissociation constant of 2 x 10(-8) M with negative cooperativity, and the last one (or two) H1's were bound cooperatively with KD = 1.8 x 10(-7) M. Regulation of the pH or the concentration of some other ion may be used in vivo to alter the interactions between HMG1 and -2, H1 histone, and DNA.