DNA-cellulose Column Chromatography Research Articles

DNA-cellulose column chromatography of phosphorylated nucleolar nonhistone proteins

A salt-extraction procedure was used to isolate a nucleolar nonhistone protein fraction, containing [32P]phosphoserine, from the nucleoli of Novikoff hepatoma ascites cells. These proteins are similar in amino-acid composition to whole nuclear (chromosomal) nonhistone proteins. DNA-cellulose column chromatography showed that this fraction contains DNA-binding phosphoproteins, some of which will bind only to homologous (Novikoff) nucleolar or nuclear DNA.

Partial purification and characterization of DNA polymerases from a rous sarcoma virus-transformed rat cell line

A new DNA polymerase was partially purified from cell-free extracts of a continuous rat cell-line (XC). The XC cells had been transformed by the Prague strain of Rous sarcoma virus but did not produce infectious virus. The molecular weight of the DNA polymerase is 70 000, as estimated by glycerol gradient centrifugation and by Sephadex gel filtration. This enzyme can be distinguished from the other cellular DNA polymerases by its elution pattern on DNA-cellulose column chromatography, its molecular weight, and its primer-template specificity. The enzyme has some characteristics of the murine leukemia virus reverse transcriptase. It is partially inhibited by immunoglobulin G purified from rabbit antiserum prepared against Rauscher leukemia virus reverse transcriptase, but is not inhibited by IgG from rat antiserum prepared against avian myeloblastosis virus reverse transcriptase. However, the XC cell enzyme can be distinguished from the murine leukemia virus reverse transcriptase by its inefficiency in copying an oligo(dG) 12 · poly(rC) primer-template.

Modification of RNA polymerase after T3 phage infection of Escherichia coli B.

E. coli B cells infected with T3 phage contain a modified host RNA polymerase in addition to the normal RNA polymerase found in uninfected cells. The modified RNA polymerase behaves differently in its elution properties from the normal enzyme on DEAE-cellulose, phosphocellulose, and DNA-cellulose column chromatography. The modified enzyme also differs from the normal polymerase in some of its enzymatic parameters. The specific activity of the modified RNA polymerase is markedly lower (i.e., (1/4)) than that of the normal enzyme. The decrease in activity is probably due to an alteration in the beta' subunit of the polymerase. A similar modification is also observed in nonpermissive cells infected with a gene-l amber mutant incapable of producing active T3 specific polymerase. An analogous modification in host RNA polymerase does not seem to occur in E. coli cells infected with T7 phage.