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Title: Ribosome synthesis during the cell cycle of Rhodomicrobium vannielii
Author: Oakley, Christopher John
ISNI:       0000 0001 3454 6683
Awarding Body: University of Warwick
Current Institution: University of Warwick
Date of Award: 1986
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A method for the purification of R. vannielii ribosomes was developed. The sizes of the two major rRNA's of R.vannielii were shown to be comparable to the 23S and 16S rRNA's of E. coli though the ribosomes themselves were smaller with subunit of sizes 46S and 26S. 32p-orthophosphate pulse labelling showed that rRNA synthesis in R. vannielii occured at a constant rate throughout a four hour period of swarmer cell maturation and reproduction. However, due to the failure of dark inhibited swarmer cells (grown photoheterotrophically under anaerobic conditions) to incorporate any of the several nucleotide precursors tested, examination of the rate of rRNA synthesis for this cell could not be carried out by this approach. In order to circumvent possible variations in membrane permeability, a method was developed to isolate membrane associated nucleoids from R. vannielii. In vitro transcriptional run off from these nucleoids was achieved but did not prove to be reproducible. Dark inhibited swarmer cells were shown to be competent in 35S-methionine uptake. Pulse labelling of r-proteins with this radiolabelled amino acid showed that r—protein synthesis, though reduced in the inhibited swarmer cell in comparison with its differentiating counterpart, does occur. This result in a cell that exhibits no increase in biomass implies that ribosome turnover occurs. Ribosomal RNA sequences were mapped on Southern blotted restriction digests of R. vannielii genomic DNA. The results indicate that this organism has at least two conventional eubacterial rRNA operons on its chromosome, though other atypical ribosomal sequences were found. The significance of these results is discussed. Two R. vannielii rDNA clones were isolated and characterized: pCO1 and pCO2 are 8.8 and 9.1 kb respectively and each contains a single 16S rRNA coding sequence. The closest linkage that these 16S genes can have to another rRNA gene is 2 Kb. This result further supports the proposal that R. vannielii possesses atypical ribosomal sequences. Three possible mechanisms by which ribosome synthesis may be influenced in the dark inhibited swarmer were investigated. No evidence was found for the action of a stringent response, DNA rearrangements or variations in DNA methylation.
Supervisor: Not available Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID:  DOI: Not available
Keywords: QH Natural history ; QP Physiology ; QR Microbiology