Interaction of deoxyribonucleic acid and histones

Abstract

Reconstitution in vitro of complexes between DNA and histone fractions (F1 and F3), each properly purified, is performed at high salt concentrations (∼ 1 M), followed by a progressive dialysis of the mixtures against decreasing ionic strengths.In a first approach, we study at pH = 5.4, the formation of a coacervate inside these mixtures, using low speed centrifugation, and measuring DNA and histone concentrations in the supernatants, as a function of the ratio of histone/DNA (w/w) in the initial mixture, and the molarity of the salt (NaCl). Interactions between DNA and histones are essentially electrostatic. However, the total number of positive charges on the histone molecules cannot explain the greater efficiency of F1 to precipitate DNA compared with F3. Uneven distribution of positive charges along histone molecules are to be considered, together with hydrophobic interactions between histones.Then by means of appropriate sucrose gradients, the existence of two complexes can be clearly demonstrated, the relative amount of each depending upon the ionic strength and the initial ratio histone/DNA in the mixture. The first complex, called “soluble”, is obtained alone only for very small values of histone/DNA ratio. With increasing amounts of histone, there appears a second complex called “aggregate”, which is in fact the first step in the formation of the coacervate.Distinct physico‐chemical factors govern each step of the binding process and it must be pointed out that the first step (soluble complex) only can be used to characterize the affinity of histone to DNA.The second step (coacervate) appears as an insoluble precipitate in the case of F1. In the case of F3, an intermediary state in the formation of the coacervate consists of stable aggregates, whose size and polydispersity are related to the initial F3/DNA ratio.The former case is somewhat similar to the complexes of DNA with polylysine, but the latter one resembles the artificial complexes between lysozyme and DNA.A tentative model is proposed for F3‐DNA aggregates where DNA is first folded into rodlike particles which in turn are organized into quasi‐spherical and highly hydrated micelles.