Project description:Division and degradation of bacterial cell walls requires coordinated action of a myriad of enzymes. This particularly applies to the elaborate cell walls of acid-fast organisms such as Mycobacterium tuberculosis, which consist of a multi-layered cell wall that contains an unusual glycan called arabinogalactan. Enzymes that cleave the D-arabinan core of this structure have not previously been identified in any organism. We have interrogated the diverse carbohydrate degrading enzymes expressed by the human gut microbiota and uncovered four families of glycoside hydrolases with activity against the D-arabinan or D-galactan components of arabinogalactan. Using novel exo-D-galactofuranosidases from gut bacteria we generated enriched D-arabinan and used it to identify Dysgonomonas gadei as a D-arabinan degrader. This enabled the discovery of endo- and exo- acting enzymes that cleave D-arabinan. We have identified new members of the DUF2961 family (GH172), and a novel family of glycoside hydrolases (DUF4185) that display endo-ᴅ-arabinofuranase activity. The DUF4185 enzymes are conserved in mycobacteria and found in many microbes, suggesting that the ability to cleave these mycobacterial glycans plays an important role in the biology of diverse organisms. All mycobacteria encode two conserved endo-D-arabinanases that display different preferences for the D-arabinan-containing cell wall components arabinogalactan and lipoarabinomannan, suggesting they are important for cell wall modification and/or degradation. The discovery of these enzymes will support future studies into the structure and function of the mycobacterial cell wall.
Project description:This study utilized the thermophile Thermus scotoductus SA-01 to determine changes in gene expression based on growth with convective or dielectric microwave heating (2.45 GHz) at 65°C. A total of 344 genes were identified as significantly differentially regulated across the three time points. Our results reveal that a large number of genes involved in cell wall recycling and biogenesis were induced, corroborating the elongated cell morphology observed in cells grown with dielectric heating. Genes associated with sugar metabolism were induced across multiple time points, while genes involved in denitrification were repressed, suggesting the different modes of heating influence metabolism. Additionally, an unexpected link was observed with regards to enzymes in which molybdenum serves as a co-factor.
Project description:A multi-layered structure known as the cell envelope separates the controlled interior of Gram-negative bacteria from a fluctuating physical and chemical environment. Transcription of genes that determine cell envelope structure and function is commonly controlled by a class of environmental regulators known as two-component signal transduction systems (TCS), which are comprised of 1) sensor histidine kinases and 2) response regulators. To discover TCS genes that contribute to cell envelope function in the intracellular mammalian pathogen, Brucella ovis, we subjected a comprehensive collection of non-essential TCS mutants to compounds that disrupt cell membranes and the peptidoglycan cell wall. Our screen led to the discovery of three TCS proteins with unusual regulatory properties that coordinately function to confer resistance to cell envelope stress and to support B. ovis replication in the intracellular niche. This tripartite regulatory system consists of the conserved cell envelope regulator, CenR, and a previously uncharacterized TCS, EssRS. The CenR and EssR response regulators bind a shared set of sites on the B. ovis chromosomes to control transcription of an overlapping set of genes with cell envelope functions. CenR directly interacts with EssR and functions to stimulate phosphoryl transfer from the EssS kinase to EssR and control steady-state levels of EssR protein in the cell via a post-transcriptional mechanism. Our data provide evidence for a new mode of TCS cross-regulation in which a non-cognate response regulator both regulates activity and influences cellular levels of a cognate TCS system.
Project description:High temperature stress leads to complex changes to plant functionality, which affects, i.a., the cell wall structure and the cell wall protein composition. In this study, the qualitative and quantitative changes in the cell wall proteome of Brachypodium distachyon leaves in response to high (40 °C) temperature stress were characterised. Using a proteomic analysis, 1533 non-redundant proteins were identified from which 338 cell wall proteins were distinguished. At a high temperature, we identified 46 differentially abundant proteins, and of these, 4 were over-accumulated and 42 were under-accumulated. The most significant changes were observed in the proteins acting on the cell wall polysaccharides, specifically, 2 over- and 12 under-accumulated proteins. Based on the qualitative analysis, one cell wall protein was identified that was uniquely present at 40 °C but was absent in the control and 24 proteins that were present in the control but were absent at 40 °C. Overall, the changes in the cell wall proteome at 40 °C suggest a lower protease activity, lignification and an expansion of the cell wall. These results offer a new insight into the changes in the cell wall proteome in response to high temperature.
2021-04-28 | MSV000087323 | MassIVE
Project description:An abundant tick sugar helps shape the unusual peptidoglycan cell-wall of Borrelia burgdorferi, the Lyme disease spirochete