Kohlstaedt L A, Cole R D
Department of Molecular and Cell Biology, University of California, Berkeley 94720.
Biochemistry. 1994 Jan 18;33(2):570-5. doi: 10.1021/bi00168a023.
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.
高迁移率族蛋白HMG1和-2以及组蛋白H1是染色质的结构成分。此前，我们报道HMG1与H1组蛋白相互作用，其方式可调节H1在体外凝聚DNA的能力，这表明这些蛋白质可能在体内共同作用以局部调节染色质的凝聚状态，可能影响复制和/或转录。在此我们表明，还原态（天然）HMG1在pH 6.0时作为四聚体与H1协同结合，解离常数为3.4×10⁻⁸ M，在pH 7.5时作为单体与H1结合，解离常数小于10⁻⁹ M。通过巯基氧化进行的变性对HMG1与H1组蛋白的相互作用有强烈影响，这表明HMG1的还原态对其功能至关重要。氧化态的HMG1在pH 7.5时无法结合H1，其在pH 6时的结合呈双相性；前三个（或两个）H1分子以2×10⁻⁸ M的解离常数负协同结合，最后一个（或两个）H1分子以KD = 1.8×10⁻⁷ M协同结合。体内可利用pH或其他一些离子浓度的调节来改变HMG1与-2、H1组蛋白和DNA之间的相互作用。
