Graduate Thesis Or Dissertation
 

Effect of actinophage infection on glucose metabolism in Streptomyces griseus

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  • Glucose metabolism of phage-infected Streptomyces griseus was examined by using the specific radioactivities of the amino acids from the extracted protein of the host and phage as an index of the catabolic pathways of glucose metabolism. Conventional methods for the study of phage-infected bacteria did not suit the host-actinophage system due to the filamentous nature of the host and the poor adsorption properties of the phage. The present method measured the specific activities of selected amino acids arising from glucose-1-¹⁴C and compared them to those of the amino acids synthesized from glucose-6-¹⁴C. A C6/C1 (ratio of the specific activity of the amino acid arising from glucose-6-¹⁴C to that arising from glucose-1-¹⁴C) specific activity ratio of 1.0 reflected the primary catabolism of glucose via the glycolytic pathway. A considerably higher ratio was expected if the pentose phosphate pathway constituted the major catabolic route since carbon one of glucose is lost at an early stage in the latter pathway. The specific activities were obtained on protein hydrolysates by automatic chromatographic procedures using the Beckman/Spinco Model 120B amino acid analyzer connected to a Packard Series 320E Flow Monitor. The procedure was tested on two cultures, Saccharomyces cerevisiae and Pseudomonas stutzeri whose widely divergent catabolic pathways had been already established elsewhere. The former dissimilates glucose primarily by glycolysis while the latter exhibits strong Entner-Doudoroff and pentose phosphate pathway activity. Using glutamic acid as a reference amino acid, a C6/C1 ratio of 1.13 and 16.3 were obtained for S. cerevisiae and P. stutzeri, respectively. These results indicated that the ratios could be used as a measure of the route of carbohydrate metabolism. Prior to determining the specific activity ratio for uninfected S. griseus, the route of glucose catabolism employed by this organism was ascertained by high-resolution radiorespirometry and the results indicated a predominant glycolytic activity. The pattern observed was similar to that found nine years earlier. The predominating glycolysis corresponded well to the C6/C1 ratio of 1.23 obtained for S. griseus using glutamic acid as the reference amino acid. To compare the route involved in the glucose catabolism of the S. griseus phage-infected system, the specific activity ratios for the amino acids from purified actinophage protein were determined. A complete amino acid composition (with the exception of tryptophan) was determined for this actinophage; however, for specific activity ratio determinations, only the acidic amino acids were analyzed. Again using glutamic acid for reference a ratio of 1.74 for the phage was obtained. The higher ratios obtained for phage protein as compared with host protein indicated an increased loss of C-1 after phage infection and during biosynthesis. The increased loss of C-1 in the phage infected system may have reflected a change in the metabolic patterns of the host to meet the demands for materials necessary for phage biosynthesis, primarily pentose for viral DNA. The direction of the switch in glucose dissimilation differs from that reported in the phage-infected Escherichia coli system and may be attributed to differences in the mechanisms by which the two systems meet the demand for pentose synthesis.
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