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Heterologous expression and in-vitro analysis of Streptococcus pneumoniae FtsEX divisome complex with peptidoglycan (PG) hydrolase PcsB and actin homologue FtsA, required for PG remodelling and cell separation
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Naskar, Souvik (2019) Heterologous expression and in-vitro analysis of Streptococcus pneumoniae FtsEX divisome complex with peptidoglycan (PG) hydrolase PcsB and actin homologue FtsA, required for PG remodelling and cell separation. PhD thesis, University of Warwick.
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Official URL: http://webcat.warwick.ac.uk/record=b3736632
Abstract
Bacterial cell division is orchestrated by the divisome complex of proteins necessary for new peptidoglycan (PG) synthesis and PG remodeling during septum formation and cell separation. These proteins have homologues in both Grampositive and Gram-negative species highlighting their fundamental biological role. The complex between FtsE and FtsX is recruited to the divisome at an early stage in mid-cell division and is required in assembling further downstream divisome proteins as well as in regulating divisome activity. Specifically, it provides a membrane anchor for an extracellular hydrolase that is required for hydrolysis of PG of old cell wall material and to enable separation of daughter cells during division.
In our heterologous expression study, we observed aberrant cell division defects in Escherichia coli (Ec) cell when subject to expression of the Streptococcus pneumoniae (Sp) FtsEX mimicking the phenotype of existing antibiotics. This phenotype can be rescued co-overexpressing SpFtsEX with its cognate peptidoglycan hydrolase; PcsB that hydrolyses Escherichia coli PG required for PG remodeling during cell separation. In this study, we have demonstrated Streptococcus pneumoniae FtsEX-FtsA and FtsEX-PcsB complexes can be isolated in-vitro using nanodiscs styrene-maleic-acid-lipid-particles (SMALP), preserving their membrane lipid environment. The protein-protein interaction studies indicate SpFtsX but not SpFtsE, interacts with the essential divisome protein SpFtsA, and PcsB successfully docks with FtsEX in the SMALP disk. Negative stain electron microscopy images and initial high resolution cryo-EM trials with these complexes indicates these tools could be prerequisite for investigating mechanistic insight about their structural-functional relationship and for further inhibitor screens for these complexes.
| Item Type: | Thesis (PhD) | ||||
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| Subjects: | Q Science > QH Natural history > QH301 Biology Q Science > QH Natural history > QH426 Genetics |
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| Library of Congress Subject Headings (LCSH): | Cell division, Cell separation, Peptidoglycans -- Synthesis, Bacterial cell walls | ||||
| Official Date: | 2019 | ||||
| Dates: |
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| Institution: | University of Warwick | ||||
| Theses Department: | Warwick Medical School | ||||
| Thesis Type: | PhD | ||||
| Publication Status: | Unpublished | ||||
| Supervisor(s)/Advisor: | Roper, David I. ; Smith, Corinne J. | ||||
| Sponsors: | Warwick Medical School ; Medical Research Council | ||||
| Format of File: | |||||
| Extent: | xxiv, 306 leaves : illustrations | ||||
| Language: | eng |
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