Abstract
A new procedure is described for purifying proteins that specifically bind to DNA. DNA is entrapped in polyacrylamide gel particles which can then be used in standard column chromatographic procedures. The method was developed using Escherichia coli DNA polymerase as the test material. The crude enzyme was applied at low ionic strength and eluted at high ionic strength with a 200-fold increase in specific activity on a single passage through the column. The method is versatile and simple and is not restricted to DNA-protein systems. Any macromolecule can be entrapped in the gel particles; these can interact with other large or small molecules in the liquid phase. The gel is stable at elevated temperatures and can therefore be used in hybridization experiments.
Full text
PDFSelected References
These references are in PubMed. This may not be the complete list of references from this article.
- Alberts B. M., Amodio F. J., Jenkins M., Gutmann E. D., Ferris F. L. Studies with DNA-cellulose chromatography. I. DNA-binding proteins from Escherichia coli. Cold Spring Harb Symp Quant Biol. 1968;33:289–305. doi: 10.1101/sqb.1968.033.01.033. [DOI] [PubMed] [Google Scholar]
- BAUTZ E. K., HALL B. D. The isolation of T4-specific RNA on a DNA-cellulose column. Proc Natl Acad Sci U S A. 1962 Mar 15;48:400–408. doi: 10.1073/pnas.48.3.400. [DOI] [PMC free article] [PubMed] [Google Scholar]
- BOLTON E. T., McCARTHY B. J. A general method for the isolation of RNA complementary to DNA. Proc Natl Acad Sci U S A. 1962 Aug;48:1390–1397. doi: 10.1073/pnas.48.8.1390. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Cavalieri L. F., Carroll E. DNA polymerase: evidence for multiple molecular species. Proc Natl Acad Sci U S A. 1968 Mar;59(3):951–958. doi: 10.1073/pnas.59.3.951. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Cavalieri L. F., Deutsch J. F., Rosenberg B. H. The Molecular Weight and Aggregation of DNA. Biophys J. 1961 Mar;1(4):301–315. doi: 10.1016/s0006-3495(61)86890-2. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Litman R. M. A deoxyribonucleic acid polymerase from Micrococcus luteus (Micrococcus lysodeikticus) isolated on deoxyribonucleic acid-cellulose. J Biol Chem. 1968 Dec 10;243(23):6222–6233. [PubMed] [Google Scholar]