Highly cooperative binding to DNA by a histone‐like, sperm‐specific protein from Spisula solidissima

In sperm of the bivalve mollusc, Spisula solidissima , the predominant protein from the highly condensed chromatin shares characteristics with both protamines and somatic histones. We have examined, in vitro, the binding of this unusual protamine‐like protein (PLP) to DNA. The binding stoichiometry was estimated by DNA thermal melting and by fluorescence titration to be about 70 base pairs of DNA per PLP molecule; thus, since the overall charge on PLP is about 140, the saturated complex should be essentially charge neutral. In order to decrease problems of aggregation, PLP was labeled with fluorescein isothiocyanate (PLP‐F) to allow binding studies at very low concentrations of the complex. However, even at the lowest protein concentrations studied (2.5 n M ), aggregation was easily detected for stoichiometries near 1. The binding was therefore studied at low ratios of PLP‐F to DNA by examining the effect of increasing salt concentration on the complexes. The resulting salt‐dissociation curves were analyzed by an approach based on the exact binding isotherm of McGhee and von Hippel [(1974) J. Biol. Chem. 86 , 469; (1976) 103 , 679]. This approach allows direct simulation of the salt‐dissociation curves and can thus be used for least squares fitting of the experimental data to determine the binding parameters. The results indicate that the binding is extremely cooperative (ω ∼ 10 5 ) and involves a surprisingly small number of monovalent ions (∼ 3–5).