Record Details
A chromatographic method for fractionating histone and ribosomal protein
ScholarsArchive at Oregon State University
| Field | Value |
|---|---|
| Title | A chromatographic method for fractionating histone and ribosomal protein |
| Names |
Haley, Lawrence Robert
(creator) Morris, Roy Owen (advisor) |
| Date Issued | 1969-05-08 (iso8601) |
| Note | Graduation date: 1969 |
| Abstract | Histones are presumed to be of chromosomal origin and provide both structural and functional elements for chromatin. In like manner the ribosomal proteins are of ribosomal origin and have analogous roles. Considerable controversy over their cellular roles exists because of the inability to compare directly both groups of proteins. In order to compare definitively the histone with the ribosomal proteins: 1) the organelles from which they are derived must be prepared under conditions which eliminate cross contamination and 2) a preparative scale method must be developed to fractionate both histone and ribosomal protein under similar conditions. A chromatographic method was developed which fractionated histone and ribosomal protein simultaneously. The histone and ribosomal proteins were isolated from beef and rat liver and germinating pea roots. Histone was acid extracted from exhaustively washed chromatin prepared from purified nuclei. Ribosomal protein was extracted from purified ribosomes. Both groups of protein were adsorbed to a column of carboxymethyl cellulose: and eluted with a sodium acetate gradient (0.01 M to 0.4 M, pH 5.6) in 6 M urea. This method is superior to IRC-50 chromatography for the following reasons: 1) no irreversible adsorption, 2) both histones and ribosomal proteins elute under the same conditions, 3) all fractions elute with a single uninterrupted gradient, 4) the reagents are more conveniently prepared and 5) the arginine-rich histones can be resolved into fractions III and IV, Beef liver and pea root histones were fractionated into six peaks corresponding in appearance to the fractions Ia, Ib, Ha, IIb, IIc and III - IV of conventional IRC-50 chromatography. Rat liver histone lacked both Ib and IIc, but peak III was resolved from peak IV with a 400 ml gradient. Ribosomal proteins were fractionated into 13 peaks (rat liver), 22 peaks (beef liver) and 17 peaks (pea root) in agreement with the number obtained from polyacrylamide gel elctrophoresis. When assayed turbidimetrically some of the histones and ribosomal proteins exhibited chromatographic identity, A spectrofluorometric assay indicated differences between chromatographically identical histone and ribosomal protein fractions. Histones and ribosomal proteins, while possessing similar physical and chemical properties, appear to have unique cellular origins and distinct roles. |
| Genre | Thesis/Dissertation |
| Topic | Histones |
| Identifier | http://hdl.handle.net/1957/46551 |
