A polyethylene glycol/dextran procedure for the isolation of chromatin proteins (histones and nonhistones) from wheat germ.
A new procedure is described for the isolation of both histone and non-histone chromatin proteins, based on a polyethylene glycol (PEG)/dextran two-phase partition system. Chromatin is solubilized in high salt (5 M NaC1) and mixed with PEG and dextran to separate proteins (partitioned into the upper, PEG-rich phase) from nucleic acid (DNA recovered almost exclusively in the lower, dextran-rich phase). The proteins are then absorbed onto Bio-Rex 70 by dialysis to low salt (0.05 M NaC1), followed by salt elution to recover first non-histone proteins (less than 0.55 M NaC1), then histones (2 M NaC1). Cross-contamination is not detectable in either group of proteins. The procedure is rapid, gentle, and lends itself well to scale-up. The proteins are kept in pH range 7.0--8.1, and are not exposed to the denaturing reagents characteristic of most preparative procedures for chromatin proteins. Though developed specifically for the isolation of proteins from chromatin of wheat germ, the procedure should be readily applicable to other sources as well. Wheat germ (isolated wheat embryos) appears to be an excellent source of chromatin proteins; 100 g yields 225--300 mg histones and 30--45 mg nonhistone proteins, depending on technique.
D000596:Amino Acids / Q000032:analysis; D002843:Chromatin / Q000378:metabolism*; D002868:Chromosomal Proteins, Non-Histone / Q000302:isolation & purification*; D012321:DNA-Directed RNA Polymerases / Q000378:metabolism; D003911:Dextrans; D006657:Histones / Q000302:isolation & purification*; D008722:Methods; D008970:Molecular Weight; D009994:Osmolar Concentration; D010944:Plants / Q000378:metabolism; D011092:Polyethylene Glycols; D014908:Triticum / Q000378:metabolism
D000596:Amino Acids; D002843:Chromatin; D002868:Chromosomal Proteins, Non-Histone; D003911:Dextrans; D006657:Histones; D011092:Polyethylene Glycols; D012321:DNA-Directed RNA Polymerases