1. Mutants of E. coli K12 devoid of acetate kinase and phosphotrans-acetylase activities have been isolated by selection for resistance to fluoroacetate. The properties of the mutants have been studied and the role of the acetate kinase/phosphotransacetylase system clarified. It has a role in acetate excretion and its control properties are consistent with a role in fermentative metabolism. It also has a role in acetate activation at high concentrations of acetate. 2. Studies of acetate incorporation by acetate kinase and phospho-transacetylase-negative mutants of E. coli K12 have revealed the existence of a second system capable of acetate activation in this organism. This system hats properties consistent with a role in the activation of acetate at low concentrations. It is saturated at 2 mM acetate and is also subject to repression by glucose. An acetate thiokinase activity could be detected in cell-free extracts and a number of its properties could be correlated with in vivo observations on the second system. The activity was induced by acetate and repressed by glucose. It had a low Km for acetate. The thiokinase also shows propionate thiokinase activity. Attempts to isolate mutants modified in this activity proved unsuccessful. 3. Problems relating to the study of monocarboxylic acid permeation have been reviewed. The uptake of acetate and lactate by washed cell suspensions of E. coli K12 has been investigated. The data obtained have been considered in the light of criteria required to establish the existence of specific transport systems. Two acetate uptake processes have been found. One shows saturation kinetics with a Km of approx. 10-5 M and its activity is repressed by glucose. The other operating at high acetate concentrations is apparently non-saturable. The effects of a number of factors on acetate uptake were investigated. Preliminary evidence that, under certain conditions, a membrane transport process may be rate-limiting in acetate uptake was obtained. The existence of specific transport systems for D- and L-lactate was established. The properties of these systems were investigated. Lactate uptake was found to be induced by lactate and repressed by glucose. Lactate was not actively concentrated within the cell, but uncoupling agents prevented its transport. The properties of a mutant defective in catabolite repression of L-lactate utilization are described. 4. Mutants of A. aerogenes 1033 devoid of acetate kinase and phospho-transacetylase activities have been isolated by selection for resistance to fluoroacetate. Their properties have been compared with those of the E. coli mutants. The acetate kinase/phosphotransacetylase system has an amphibolic role in A. aerogenes. No evidence for the operation of an alternative mechanism of acetate activation was obtained. The coarse control properties of the acetate kinase/phosphotransacetylase system were studied under a variety of conditions. The role of acetic acid as the inducer of the enzymes of the butanediol-forming system of A. aerogenes was established. Possible relationships between the operation of the acetate excreting system and the operation of the butanediol-forming system are discussed.
| Identifer | oai:union.ndltd.org:bl.uk/oai:ethos.bl.uk:674061 |
| Date | January 1972 |
| Creators | Brown, Thomas David Kay |
| Publisher | University of Leicester |
| Source Sets | Ethos UK |
| Detected Language | English |
| Type | Electronic Thesis or Dissertation |
| Source | http://hdl.handle.net/2381/35184 |
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