Solid-state NMR studies of peptidoglycan and teichoic acids in the bacterial cell wall and bacterial biofilms
Abstract/Contents
- Abstract
- Gram-positive bacteria surround themselves with a cell wall that is essential to cell survival and serves as a major target for antibiotics. The cell wall is composed primarily of two polymeric macromolecules: peptidoglycan (PG) and teichoic acid (TA). During infection, bacteria can further elaborate a multi-component extracellular matrix beyond the cell wall to form a biofilm community, enmeshing many cells, that improves adhesion, increases virulence, and helps avoid host immune responses. Cell walls and biofilm matrices are insoluble and heterogeneous materials that pose a challenge to analysis with biophysical techniques like X-ray crystallography, solution NMR, and mass spectrometry. We are implementing solid-state NMR methods to access atomic-level compositional parameters of cell walls and the extracellular matrix, particularly doing so in the context of intact biofilms to address questions that cannot be answered by attempted digestions and solution-based analysis. I will report on our new approach to quantify the relative proportions of PG and TA in isolated cell walls through 13C analysis, and how we have extended this approach to intact whole cells and protoplasts to identify several characteristic spectral features of cell walls. These allow us to gauge cell wall levels quantitatively. Further 15N analysis revealed a uniquely shift-resolved peak in whole cells entirely attributed to TA D-alanylation. We use these markers alongside isotopic-labeling experiments to characterize the distinct mode of actions of a panel of cell wall synthesis inhibitors. Through our analysis, we also found that cells in stationary phase exhibit reduced levels of TA with increased PG. We have now discovered that TA D-alanylation decreases significantly in cells that make a robust extracellular matrix, as visualized by EM. Similarly, cells treated with a small molecule inhibitor of teichoic acid synthesis appear to make more developed biofilms. Our solid-state NMR measurements are integrated with proteomic and microscopic analysis of biofilm cells and reveal direct composition-function relationships. Together, our results demonstrate that S. aureus PG and TA production are tunable, and that decreased TA production or modification occurs as cells form biofilms.
Description
| Type of resource | text |
|---|---|
| Form | electronic resource; remote; computer; online resource |
| Extent | 1 online resource. |
| Place | California |
| Place | [Stanford, California] |
| Publisher | [Stanford University] |
| Copyright date | 2019; ©2019 |
| Publication date | 2019; 2019 |
| Issuance | monographic |
| Language | English |
Creators/Contributors
| Author | Romaniuk, Joseph Anthony |
|---|---|
| Degree supervisor | Cegelski, Lynette |
| Thesis advisor | Cegelski, Lynette |
| Thesis advisor | Boxer, Steven G. (Steven George), 1947- |
| Thesis advisor | Solomon, Edward I |
| Degree committee member | Boxer, Steven G. (Steven George), 1947- |
| Degree committee member | Solomon, Edward I |
| Associated with | Stanford University, Department of Chemistry. |
Subjects
| Genre | Theses |
|---|---|
| Genre | Text |
Bibliographic information
| Statement of responsibility | Joseph A. H. Romaniuk. |
|---|---|
| Note | Submitted to the Department of Chemistry. |
| Thesis | Thesis Ph.D. Stanford University 2019. |
| Location | electronic resource |
Access conditions
- Copyright
- © 2019 by Joseph Anthony Romaniuk
- License
- This work is licensed under a Creative Commons Attribution Non Commercial 3.0 Unported license (CC BY-NC).
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