Field, J., Nikawa, J., Broek, D., MacDonald, B., Rodgers, L., Wilson, I. A., Lerner, R. A., Wigler, M. H. (May 1988) Purification of a RAS-responsive adenylyl cyclase complex from Saccharomyces cerevisiae by use of an epitope addition method. Molecular & Cellular Biology, 8 (5). pp. 2159-65. ISSN 0270-7306
Abstract
We developed a method for immunoaffinity purification of Saccharomyces cerevisiae adenylyl cyclase based on creating a fusion with a small peptide epitope. Using oligonucleotide technology to encode the peptide epitope we constructed a plasmid that expressed the fusion protein from the S. cerevisiae alcohol dehydrogenase promoter ADH1. A monoclonal antibody previously raised against the peptide was used to purify adenylyl cyclase by affinity chromatography. The purified enzyme appeared to be a multisubunit complex consisting of the 200-kilodalton adenylyl cyclase fusion protein and an unidentified 70-kilodalton protein. The purified protein could be activated by RAS proteins. Activation had an absolute requirement for a guanine nucleoside triphosphate.
| Item Type: | Paper |
|---|---|
| Additional Information: | |
| Uncontrolled Keywords: | Adenylate Cyclase isolation & purification Amino Acid Sequence Antibodies Monoclonal immunology Chromatography Affinity Epitopes immunology Fungal Proteins isolation & purification physiology Guanine Nucleotides pharmacology Molecular Sequence Data Peptides immunology Recombinant Fusion Proteins immunology isolation & purification Saccharomyces cerevisiae enzymology ras Proteins |
| Subjects: | bioinformatics > genomics and proteomics > genetics & nucleic acid processing > DNA, RNA structure, function, modification > genes, structure and function > genes: types > RAS organism description > yeast > Saccharomyces biotechnology > chromatography > affinity chromatography biotechnology > chromatography bioinformatics > genomics and proteomics > genetics & nucleic acid processing > DNA, RNA structure, function, modification > genes, structure and function > genes: types > oncogene |
| CSHL Authors: | |
| Communities: | CSHL labs > Wigler lab |
| Depositing User: | CSHL Librarian |
| Date: | May 1988 |
| Date Deposited: | 13 Apr 2012 20:07 |
| Last Modified: | 04 Nov 2016 20:46 |
| PMCID: | PMC363397 |
| Related URLs: | |
| URI: | https://repository.cshl.edu/id/eprint/26181 |
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