Rapid purification of native C protein from nuclear ribonucleoprotein particles.
A rapid three step procedure is described for the purification of C protein from HeLa 40 S hnRNP particles. The procedure takes advantage of the salt resistant RNA binding of C protein, the size of the C protein-RNA complex, and the strong binding of C protein to an anion-exchange resin. Typically 120 micrograms of C protein is obtained from 4.0 X 10(9) cells with greater than 95% electrophoretic purity. Proteins C1 and C2 copurify in the ratio of 3.5 Cl to 1 C2. The purified C protein participates in hnRNP particle reconstitution and on this basis is judged to be native. The purified C protein binds to a gel filtration matrix at 0.5 M NaCl but at higher salt concentrations it elutes before the marker protein, apoferritin (Mr = 443,000). An abbreviated two step purification procedure utilizing anion-exchange chromatography is also described. This procedure results in relatively pure C protein, as well as a useful separation of the other hnRNP proteins.
Barnett
S F
SF
Department of Molecular Biology, Vanderbilt University, Nashville, TN 37235.
LeStourgeon
W M
WM
Friedman
D L
DL
D002352:Carrier Proteins / Q000302:isolation & purification* / Q000378:metabolism; D002467:Cell Nucleus / Q000032:analysis*; D002850:Chromatography, Gel; D002852:Chromatography, Ion Exchange; D004591:Electrophoresis, Polyacrylamide Gel; D006367:HeLa Cells; D006801:Humans; D012313:RNA / Q000378:metabolism; D012261:Ribonucleoproteins / Q000032:analysis*; D012965:Sodium Chloride
D002352:Carrier Proteins; D012261:Ribonucleoproteins; C020723:citrate-binding transport protein; D012965:Sodium Chloride; D012313:RNA