Purification of the phosphoroclastic system of Escherichia coli was undertaken. A 4-fold purification of the crude extract was obtained using heat precipitation and protamine sulfate. Some purification of Knappe's A fraction was obtained with ammonium sulfate and acetone. Evidence was obtained for the existence of two factors in the A fraction. Other purification techniques gave little success. Good evidence was obtained for the involvement of phosphotrans-acetylase in the reaction sequence. Phosphotransacetylase was purified 750-fold and characterized. The reversibility of the reaction was studied with carbon 14. Formate fixed readily into pyruvate only when pyruvate was present. Acetyl coenzyme A fixed into pyruvate also but to a much smaller degree. Better fixation without pyruvate was obtained after prior incubation and consumption of pyruvate. The dilution effect, inhibitors, effect of light, and a possible role of coenzyme B_12 were investigated. By using a deuterium label, it was shown that a hydrogen does not shift directly from the methyl group of pyruvate to formate in the reaction mechanism.
Identifer | oai:union.ndltd.org:BGMYU2/oai:scholarsarchive.byu.edu:etd-9386 |
Date | 01 May 1970 |
Creators | Winkel, Cleve R. |
Publisher | BYU ScholarsArchive |
Source Sets | Brigham Young University |
Detected Language | English |
Type | text |
Source | Theses and Dissertations |
Rights | http://lib.byu.edu/about/copyright/ |
Page generated in 0.0019 seconds