Analysis of the most tightly bound proteins in eukaryotic DNA.
DNA isolated by procedures generally considered to be most efficient for purifying DNA still contains detectable peptide components. The characteristics of this material and the stability of its linkage to DNA were investigated: DNA released from [35S]methionine-labelled cells by SDS in the presence of proteases contains a significant amount of 35S label which is not removed by additional treatment with proteases and phenol and which cosediments and cobands together with DNA on alkaline gradients. Furthermore, some peptide material which is copurified with native DNA and which remains complexed with DNA after alkali treatment can be labelled with 125I and analyzed on SDS-polyacrylamide-gels. The amino acid analysis of hydrolysates of purified DNA gives a rough estimate of the amount of the peptide material which is copurified with DNA. The results indicate that distinct proteins between 54 000 and 68 000 daltons in size are not removed from DNA by phenol, proteases, alkali or by any combination of these treatments. They can only be isolated by degradation of DNA. This extreme stability of the DNA-protein linkage indicates that these proteins are not merely contaminants which are difficult to eliminate but are rather covalently or otherwise bound (alkali-stable) to DNA. The size of these proteins and the stability of their linkage to DNA suggests that they are related to the class of non-histone proteins which are thought to be involved in chromatin structure e.g. by keeping DNA in a supercoiled state. Other possible functions are discussed.
D000596:Amino Acids / Q000032:analysis; D000818:Animals; D002286:Carcinoma, Ehrlich Tumor / Q000032:analysis*; D004273:DNA, Neoplasm / Q000032:analysis / Q000302:isolation & purification; D003852:Deoxyribonucleoproteins / Q000032:analysis*; D051379:Mice; D008970:Molecular Weight; D009698:Nucleoproteins / Q000032:analysis*; D011485:Protein Binding
D000596:Amino Acids; D004273:DNA, Neoplasm; D003852:Deoxyribonucleoproteins; D009698:Nucleoproteins