Investigations on cytochrome bd from Escherichia coli and Mycobacterium tuberculosis.
In recent years, bacterial respiratory chains have attracted attention, in particular regarding the role of respiratory proteins in pathogenicity or as target of new antibiotics. The majority of next-generation drugs for treatment of tuberculosis target proteins in the mycobacterial respiratory chain. Respiratory chain proteins are also under investigation in various other bacterial pathogens, where this central pathway can be important for survival in the human host. Cytochrome bd is a terminal oxidase found in the respiratory chains of many bacteria. Cytochrome bd is necessary in pathogenic E. coli for host colonization, whereas in Mycobacterium tuberculosis this protein is seen as promising antibiotic target. Despite of intense research on cytochrome bd, no drugs targeting this protein have yet been approved. Improved insight into structure and function of cytochrome bd may help to develop effective drugs against this important respiratory protein. In this thesis, cytochrome bd from Escherichia coli and from Mycobacterium tuberculosis is investigated. Biochemical approaches showed that an insert in the quinone binding site (Q-loop) is needed for full assembly/functionality of cytochrome bd from E. coli. In contrast, a recently discovered new subunit of this enzyme surprisingly appeared to be not necessary for functionality under the conditions tested. Interestingly, a second isoform of cytochrome bd in E. coli differed from the first isoform in terms of subunit composition, enzymatic activity and regulation of activity, providing hints for why some bacteria, like E. coli, use more than one cytochrome bd isoform. In the final chapter of this thesis, cytochrome bd from Mycobacterium tuberculosis was investigated. We demonstrated that the functionality of this protein can be monitored in intact M. tuberculosis bacteria using a convenient electrode system. This experimental system was applied to determine the impact of prominent respiratory chain inhibitors on respiration by living Mycobacterium tuberculosis cells.
https://research.vu.nl/ws/files/168173901/H%20%20Ghasemi%20Goojani%20-%20toc.pdf
https://research.vu.nl/ws/files/168173903/H%20%20Ghasemi%20Goojani%20-%20title_page.pdf
https://research.vu.nl/ws/files/168173905/H%20%20Ghasemi%20Goojani%20-%20thesis_redacted.pdf