Title

Purification and mechanism studies on the phosphoroclastic reaction of escherichia coli

Abstract

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.

Degree

PhD

College and Department

Chemistry and Biochemistry

Rights

http://lib.byu.edu/about/copyright/

Date Submitted

1970-05-01

Document Type

Dissertation

Handle

http://hdl.lib.byu.edu/1877/Letd693

Keywords

Enzymes, Escherichia coli.

Language

English

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