DIFFERENTIAL SILVER-STAINING SODIUM DODECYL SULFATE-POLYACRYLAMIDE GEL-ELECTROPHORESIS - A NONISOTOPIC METHOD FOR CHARACTERIZING GEL-SEPARATED HISTONE-DNA COMPLEXES

Some nonspecific, DNA-binding proteins, like the linker histones, prec ipitate DNA upon binding. This is a poorly understood process that Lim its analysis of such nucleoprotein complexes using standard gel electr ophoresis, To circumvent this problem, low concentrations of glutarald ehyde were used to crosslink the Linker histones to DNA; then the part ially crosslinked complexes were solubilized in SDS2 and separated by SDS-PAGE. Differential detection was accomplished using two different silver staining protocols that preferentially stained either proteins or nucleic acids. A technique was developed which allows the relative proportion of linker histones and DNAs in each detected band to be det ermined, and is referred to as differential staining SDS-PAGE (DS-SDS PAGE). DS-SDS-PAGE provides a novel, nonisotopic means for characteriz ing multiple nucleoprotein bands separated by polyacrylamide gel elect rophoresis. In applying this method to a model linker histone-DNA stud y, we were able to detect both protein-DNA and protein-protein contact s that are important in linker histone assembly onto DNA (C) 1998 Acad emic Press.