| 1. |
- Bratanis, Eleni, et al.
(författare)
-
Biotechnological Potential of Bdellovibrio and Like Organisms and Their Secreted Enzymes
- 2020
-
Ingår i: Frontiers in Microbiology. - : Frontiers Media SA. - 1664-302X. ; 11
-
Forskningsöversikt (refereegranskat)abstract
- Bdellovibrio and like organisms (BALOs) are obligate predatory bacteria that selectively prey on a broad range of Gram-negative bacteria, including multidrug-resistant human pathogens. Due to their unique lifestyle, they have been long recognized as a potential therapeutic and biocontrol agent. Research on BALOs has rapidly grown over the recent decade, resulting in many publications concerning molecular details of bacterial predation as well as applications thereof in medicine and biotechnology. This review summarizes the current knowledge on biotechnological potential of obligate predatory bacteria and their secreted enzymes.
|
|
| 2. |
- Bukowska-Faniband, Ewa, et al.
(författare)
-
Cortex synthesis during Bacillus subtilis sporulation depends on the transpeptidase activity of SpoVD.
- 2013
-
Ingår i: FEMS Microbiology Letters. - : Oxford University Press (OUP). - 1574-6968 .- 0378-1097. ; 346:1, s. 65-72
-
Tidskriftsartikel (refereegranskat)abstract
- The nonessential process of peptidoglycan synthesis during Bacillus subtilis sporulation is one model to study bacterial cell wall biogenesis. SpoVD is a class B high-molecular weight penicillin-binding protein that is specific for sporulation. Strains lacking this protein produce spores without the peptidoglycan cortex layer and are heat-sensitive. The detailed functions of the four different protein domains of the SpoVD protein are unknown and the observed phenotype of strains lacking the entire protein could be an indirect defect. We therefore inactivated the transpeptidase domain by substitution of the active site serine residue. Our results demonstrate that endospore cortex synthesis depends on the transpeptidase activity of SpoVD specifically. This article is protected by copyright. All rights reserved.
|
|
| 3. |
- Bukowska-Faniband, Ewa
(författare)
-
On the role of penicillin-binding protein SpoVD in endospore cortex assembly
- 2015
-
Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- Bacteria of the genera Bacillus and Clostridium can form endospores as a strategy to survive unfavourable environmental conditions. Endospore formation involves synthesis of cortex, a thick layer of modified peptidoglycan that surrounds the spore. This layer is required for heat resistance of the spore and mutant spores lacking the cortex layer can be identified by a simple heat shock assay. B. subtilis SpoVD is a class B, high molecular weight, penicillin-binding protein (PBP) essential for spore cortex peptidoglycan synthesis. The exact role of the protein in cortex assembly is nknown but it most likely catalyses the formation of cross-links between glycan strands in nascent peptidoglycan. SpoVD deficient strains produce heat sensitive spores without cortex layer. Two conserved cysteine residues (Cys332 and Cys351) in the transpeptidase domain of SpoVD seem important for activity of the enzyme. They can form an intramolecular disulfide bond and this is catalysed by the membrane-bound thiol-disulfide oxidoreductase BdbD. The disulfide bond in SpoVD is located close to the transpeptidase active site and blocks the function of the protein. The bond is broken by the action of StoA, a sporulation-specific membrane-bound thiol-disulfide oxidoreductase. Based on these findings a thiol-based redox switch regulation of SpoVD activity was proposed in 2010. The aim of this PhD project was to elucidate the function of SpoVD in cortex synthesis and to find out the physiological role of the proposed switch and the two cysteine residues in SpoVD. In depth investigation of the process of cortex assembly contribute to our understanding of peptidoglycan synthesis in general. This is of considerable medicinal interest since, e.g., bacterial cell wall synthesis is an effective target for many antibiotics in clinical use, such as penicillins and cephalosporins and eventual new drugs. I demonstrate, by the use of a constructed SpoVD active site mutant strain, that synthesis of cortex explicitly depends on the transpeptidase activity of the protein. I show that the C-terminal PASTA domain of SpoVD is not important for the function of the protein in cortex synthesis. My results from in vitro experiments with several isolated protein variants strengthen the view that SpoVD is a specific target for StoA. My findings, supported by data available in the literature, indicate that the two cysteine residues in SpoVD affect the dynamics of the transpeptidase domain. Finally, I propose a revised model for the function of BdbD and StoA in modulation of the redox state of SpoVD, where BdbD and StoA are suggested to act (mainly) during the folding of newly synthesised SpoVD.
|
|
| 4. |
- Bukowska-Faniband, Ewa, et al.
(författare)
-
Studies on Bd0934 and Bd3507, two secreted nucleases from bdellovibrio bacteriovorus, reveal sequential release of nucleases during the predatory cycle
- 2020
-
Ingår i: Journal of Bacteriology. - 0021-9193. ; 202:18
-
Tidskriftsartikel (refereegranskat)abstract
- Bdellovibrio bacteriovorus is an obligate predatory bacterium that invades and kills a broad range of Gram-negative prey cells, including human pathogens. Its potential therapeutic application has been the subject of increased research interest in recent years. However, an improved understanding of the fundamental molecular aspects of the predatory life cycle is crucial for developing this bacterium as a “living antibiotic.” During intracellular growth, B. bacteriovorus secretes an arsenal of hydrolases, which digest the content of the host cell to provide growth nutrients for the predator, e.g., prey DNA is completely degraded by the nucleases. Here, we have, on a genetic and molecular level, characterized two secreted DNases from B. bacteriovorus, Bd0934 and Bd3507, and determined the temporal expression profile of other putative secreted nucleases. We conclude that Bd0934 and Bd3507 are likely a part of the predatosome but are not essential for the predation, host-independent growth, prey biofilm degradation, and self-biofilm formation. The detailed temporal expression analysis of genes encoding secreted nucleases revealed that these enzymes are produced in a sequential orchestrated manner. This work contributes to our understanding of the sequential breakdown of the prey nucleic acid by the nucleases secreted during the predatory life cycle of B. bacteriovorus. IMPORTANCE Antibiotic resistance is a major global concern with few available new means to combat it. From a therapeutic perspective, predatory bacteria constitute an interesting tool. They not only eliminate the pathogen but also reduce the overall pool of antibiotic resistance genes through secretion of nucleases and complete degradation of exogenous DNA. Molecular knowledge of how these secreted DNases act will give us further insight into how antibiotic resistance, and the spread thereof, can be limited through the action of predatory bacteria.
|
|
| 5. |
- Bukowska-Faniband, Ewa, et al.
(författare)
-
The PASTA domain of penicillin-binding protein SpoVD is dispensable for endospore cortex peptidoglycan assembly in Bacillus subtilis.
- 2015
-
Ingår i: Microbiology. - : Microbiology Society. - 1465-2080 .- 1350-0872. ; 161:Online 6 January, 2014, s. 330-340
-
Tidskriftsartikel (refereegranskat)abstract
- Peptidoglycan is the major structural component of the bacterial cell wall. Penicillin-binding proteins (PBPs), located at the exterior of the cytoplasmic membrane, play a major role in peptidoglycan synthesis and remodelling. A PASTA domain (penicillin-binding protein and serine/threonine kinase associated domain) of about 65 residues is found at the C-terminal end of some PBPs and eukaryotic-like protein serine/threonine kinases in a variety of bacteria. The function of PASTA domains is not understood but some of them are thought to bind uncrosslinked peptidoglycan. Bacillus subtilis has sixteen different PBPs but only two of them, Pbp2b and SpoVD, contain a PASTA domain. SpoVD is specific for sporulation and essential for endospore cortex peptidoglycan synthesis. We have studied the role of the PASTA domain in SpoVD by deleting this domain and analyzed effects on endospore formation and subcellular localization of SpoVD. Our results demonstrate that the PASTA domain in SpoVD is not essential for cortex synthesis and not important for targeting SpoVD to the forespore outer membrane during sporulation.
|
|
| 6. |
- Hederstedt, Lars, et al.
(författare)
-
Transpeptidase activity of penicillin-binding protein SpoVD in peptidoglycan synthesis conditionally depends on the disulfide reductase StoA
- 2017
-
Ingår i: Molecular Microbiology. - : Wiley. - 1365-2958 .- 0950-382X. ; 105, s. 98-114
-
Tidskriftsartikel (refereegranskat)abstract
- Endospore cortex peptidoglycan synthesis is notrequired for bacterial growth but essential for endo-spore heat resistance. It therefore constitutes anamenable system for research on peptidoglycan bio-genesis. The Bacillus subtilis sporulation-specificclass B penicillin-binding protein (PBP) SpoVD andmany homologous PBPs contain two conserved cys-teine residues of unknown function in the transpepti-dase domain – one as residue x in the SxN catalyticsite motif and the other in a flexible loop near the cat-alytic site. A disulfide bond between these residuesblocks the function of SpoVD in cortex synthesis.With a combination of experiments with purified pro-teins and B. subtilis mutant cells, it was shown thatin active SpoVD the two cysteine residues most prob-ably interact by hydrogen bonding and that this isimportant for peptidoglycan synthesis in vivo. It wasfurthermore demonstrated that the sporulation-specific thiol-disulfide oxidoreductase StoA reducesSpoVD and that requirement of StoA for cortex syn-thesis can be suppressed by two completely differenttypes of structural alterations in SpoVD. It is con-cluded that StoA plays a critical role mainly duringmaturation of SpoVD in the forespore outer mem-brane. The findings advance our understanding ofessential PBPs and redox control of extra-cytoplasmic protein disulfides in bacterial cells.
|
|
| 7. |
- Sidarta, Margareth, et al.
(författare)
-
Forespore targeting of SpoVD in Bacillus subtilis is mediated by the N-terminal part of the protein.
- 2018
-
Ingår i: Journal of Bacteriology. - 0021-9193. ; 200:e00163-18
-
Tidskriftsartikel (refereegranskat)abstract
- SpoVD and PBP4b are structurally very similar high-molecular-weight, class B penicillin-binding proteins produced early during sporulation in Bacillus subtilis. SpoVD is known to be essential for endospore cortex synthesis and thereby the production of heat-resistant spores. The role of PBP4b is still enigmatic. Both proteins are synthesized in the cytoplasm of the mother cell. PBP4b remains in the cytoplasmic membrane of the mother cell, whereas SpoVD accumulates in the forespore outer membrane. By the use of SpoVD/PBP4b chimeras with swapped protein domains, we show that the N-terminal part of SpoVD, containing the single transmembrane region, determines the forespore targeting of the protein.
|
|
