Location on the chromosome of Salmonella typhimurium of genes governing pyrimidine metabolism. II. Uridine kinase, cytosine deaminase and thymidine kinase.
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Biomedical subjects
Publications and source records attributed to J L Ingraham.
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The pathways by which uracil, cytosine, uridine, cytidine, deoxyuridine, and deoxycytidine are metabolized by Salmonella typhimurium are established. The various 5-fluoropyrimidine analogues are shown to exert their toxic effects only after having been converted to the nucleotide level, and these conversions are shown to be catalyzed by the same enzymes which similarly convert the natural substrates. Methods for isolating mutant strains blocked in various steps of metabolism of pyrimidine bases and nucleosides are described.
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The minimal temperature for growth of Escherichia coli ML30 in glucose minimal medium is between 7.5 and 7.8 C. After transfer to subminimal temperature, growth occurs but is not sustained.
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As part of a study on the effect of low temperature on cellular regulatory processes, a class of lactose-negative mutants of Escherichia coli K-12 was isolated which could use lactose as a sole carbon and energy source at 37 C, but which could not use this sugar at 20 C. The lactose operon of the mutants functioned normally at 20 C. Galactose exhibited a strong inhibitory effect on growth, especially at 20 C. Growth of the mutants on glycerol was stopped at 20 C and slowed considerably at 37 C if galactose was added to the medium. Making the mutants galactose-positive eliminated the cold sensitivity of lactose utilization. One mutant was shown to be galactose-1-phosphate uridyl transferase-negative, galactose-kinase heat-sensitive, and uridine diphosphate-galactose-4-epimerase-positive. It is postulated that the mutant is able to phosphorylate galactose at 20 C (if only at a very low rate), but lacking transferase it is poisoned by the accumulation of galactose-1-phosphate. At 37 C, galactokinase is nonfunctional and the mutant grows on the glucose moiety of lactose.
When a culture of Escherichia coli ML30 growing exponentially at 37 C in a glucose minimal medium was shifted abruptly to 10 C, growth decreased for about 4.5 hr. There was no net synthesis of deoxyribonucleic acid (DNA), ribonucleic acid (RNA), and protein. The cells, however, respired at a rate characteristic of cells growing in the steady state at 10 C and were able to accumulate alpha-methyl-d-glucoside. When growth recommenced at 10 C, protein synthesis started at 4 hr, RNA synthesis, with a burst at 6 hr, and DNA synthesis, with a burst at 7 hr. One synchronous division occurred at about 11 hr after shifting to 10 C. There was no alteration in the steady-state RNA to protein ratio. The results are discussed in relation to other reported effects of shifts in environmental conditions. The lag at 10 C was dependent on prior conditions of growth at 37 C. Growth at 37 C under conditions giving catabolite repression were necessary for the lag to be established on shifting to 10 C.
A cold-sensitive mutant of Pseudomonas putida has been isolated which grows normally at 30 C but is unable to grow on mandelate as a source of carbon at 15 C. The mutation results in the inability of the strain to carry out the reaction catalyzed by cis,cis-muconate lactonizing enzyme at low temperature and must lie in the structural gene for that enzyme, because the mutant enzyme produced at 30 C shows altered thermal stability. The mutant enzyme is not intrinsically cold-labile, nor is it cold-labile at the moment of synthesis. The activity of the mutant enzyme is not inhibited at low temperature. Evidence is presented to establish that this mutation in the structural gene coding for cis,cis-muconate lactonizing enzyme results in the lack of expression of that gene at low temperature.
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Shaw, Maxwell K. (University of California, Davis), and John L. Ingraham. Fatty acid composition of Escherichia coli as a possible controlling factor of the minimal growth temperature. J. Bacteriol. 90:141-146. 1965.-If Escherichia coli ML30 is shifted from 37 to 10 C during exponential growth in glucose minimal medium, a 4.5-hr lag results. During this lag, the proportion of unsaturated fatty acids increases in the cellular lipids. However, the adjustment of the fatty acid composition does not appear to be prerequisite to growth at 10 C. If shifts are made to 10 C into minimal medium containing glucose after starvation for glucose at 37 C for 0.5 and 16 hr, the lag periods at 10 C are 4.5 and 6 hr, respectively. Withholding glucose during the lag periods does not affect the duration of the lag periods, but no change in fatty acid composition occurs if glucose is not present. Supplementing the medium with glucose after the lag period permits immediate growth at 10 C; however, the fatty acid composition is still typical of cells grown at 37 C. It is concluded that the fatty acid composition of cells does not determine the minimal temperature of growth.
O'Donovan, Gerard A. (University of California, Davis), Catherine L. Kearney, and John L. Ingraham. Mutants of Escherichia coli with high minimal temperatures of growth. J. Bacteriol. 90:611-616. 1965.-Three general classes of mutants showing increased minimal temperatures of growth have been isolated from Escherichia coli. These mutants do not grow at temperatures below 20 C, although their parents can grow at temperatures as low as 8 C. The first class of mutants (K-I) cannot grow below 20 C in either complex or minimal medium, but grows at nearly normal rates at 37 C on both types of media. Normal growth rate at 20 C can be conferred on these mutants by infection at a low multiplicity with a transducing phage grown on the parent. The second class of mutants (K-II) fails to grow only in minimal medium at 20 C. These mutants are characterized by their singular response to specific nutrients in minimal medium at 20 C. The third class of mutants (K-III) grows normally in minimal medium at all temperatures with either glucose or glycerol as the carbon source, but does not grow at 20 C with lactose as the carbon source.
Marr, Allen G. (University of California, Davis) and John L. Ingraham. Effect of temperature on composition of fatty acids in Escherichia coli. J. Bacteriol. 84:1260-1267. 1962.-Variations in the temperature of growth and in the composition of the medium alter the proportions of individual fatty acids in the lipids of Escherichia coli. As the temperature of growth is lowered, the proportion of unsaturated fatty acids (hexadecenoic and octadecenoic acids) increases. The increase in content of unsaturated acids with a decrease in temperature of growth occurs in both minimal and complex media. Cells harvested in the stationary phase contained large amounts of cyclopropane fatty acids (methylenehexadecanoic and methylene octadecanoic acids) in comparison with cells harvested during exponential growth. Cells grown in a chemostat, limited by the concentration of ammonium salts, show a much higher content of saturated fatty acids (principally palmitic acid) than do cells harvested from an exponentially-growing batch culture in the same medium. Cells grown in a chemostat, limited by the concentration of glucose, show a slightly higher content of unsaturated fatty acids than cells from the corresponding batch culture. The results do not indicate a direct relation between fatty acid composition and minimal growth temperature.
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