Removal and Replacement of Ribosomal Proteins: Effects on Bacterial Fitness and Ribosome Function
2011 (English)Doctoral thesis, comprehensive summary (Other academic)
Protein synthesis is a complex process performed by sophisticated cellular particles known as ribosomes. Although RNA constitutes the major structural and functional component, ribosomes from all kingdoms contain an extensive array of proteins with largely undefined functional roles. The work presented in this thesis addresses ribosomal complexity using mutants of Salmonella typhimurium to examine the physiological effects of ribosomal protein (r-protein) removal and orthologous replacement on bacterial fitness and ribosome function.
The results of paper I demonstrate that removal of small subunit protein S20 conferred two independent translation initiation defects: (i) a significant reduction in the rate and extent of mRNA binding and (ii) a drastic decrease in the yield of 70S complexes caused by an impairment in subunit association. The topographical location of S20 in mature 30S subunits suggests that these perturbations are the result of improper orientation of helix 44 of the 16S rRNA when S20 is absent. In paper II we show that the major functional impairment associated with loss of large subunit protein L1 manifested as an increase in free ribosomal subunits at the expense of translationally active 70S particles. Furthermore, the formation of free ribosomal subunits was imbalanced suggesting that L1 is required to suppress degradation or promote formation of 30S subunits. Compensatory evolution revealed that mutations in other large subunit proteins mitigate the cost of L1 removal, in one case seemingly via an increase in 70S complex formation. As shown in paper III, the large fitness costs associated with complete removal of r-proteins is in contrast to the generally mild costs of orthologous protein replacement, even in the absence of a high degree of homology to the native protein. This clearly demonstrates the robustness and plasticity of the ribosome and protein synthesis in general and it also implies that functional constraints are highly conserved between these proteins. The findings of paper III also allowed us to examine the barriers that constrain horizontal gene transfer and we find that increased gene dosage of the sub-optimal heterologous protein may be an initial response to stabilize deleterious transfer events. Overall the results highlight the requirement of r-proteins for the maintenance of ribosomal structural integrity.
Place, publisher, year, edition, pages
Uppsala: Acta Universitatis Upsaliensis , 2011. , 93 p.
Digital Comprehensive Summaries of Uppsala Dissertations from the Faculty of Medicine, ISSN 1651-6206 ; 667
ribosome, protein synthesis, ribosomal proteins, translation initiation, ribosome biogenesis, fitness costs, compensatory evolution, horizontal gene transfer
Microbiology in the medical area Biochemistry and Molecular Biology
Research subject Microbiology
IdentifiersURN: urn:nbn:se:uu:diva-150401ISBN: 978-91-554-8061-5 (print)OAI: oai:DiVA.org:uu-150401DiVA: diva2:408601
2011-05-20, C8:301, BMC, Husargatan 3, Uppsala, 09:00 (English)
Buckingham, Richard, Professor
Andersson, Dan I., Professor
List of papers