Assembly of DNA with histones and nonhistone chromosomal proteins in vitro

Publisher Website
Google Scholar
Abstract
Protein binding assays, thermal denaturation, and circular dichroism were performed to examine the possible effects of histones on nonhistone chromosomal protein (NHCP) interactions with DNA. Mouse Krebs II [ascites tumor cells] chromosomal proteins were fractionated into 3 discrete fractions: M0, 5 M urea-soluble NHCP; M1, 5 M urea-1 M NaCl-soluble NHCP from 5 M urea-extracted chromatin; and M3, 5 M urea-3 M NaCl-soluble chromosomal proteins from 5 M urea-1 M NaCl-extracted chromatin. These fractions contain heterogeneous populations of NHCP, and differentially affected histone binding to DNA by methods of reconstitution, or by direct binding of M0, M1 or M3 to urea-salt reconstituted DNA with histones. M0 exerted a unique effect on the thermal denaturation and circular dichroic spectra of DNA-histone complexes. M0 from Krebs II chromatin also competed for DNA sites in the presence of M0 from mouse liver chromatin. In addition, in 5 M urea, pH 8.0, histone binding to DNA reached saturation at 1.85 mg/mg of DNA, higher than the in vivo ratio of 1.00 mg/mg of DNA. Saturation of histone binding to DNA occurred only in the presence of 5 M urea, resulting in a reduction of nonspecific histone-histone interactions on DNA.