Polyketide synthase-derived sphingolipids determine microbiota-mediated protection against pathogens inC. elegans
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PROVIDER: PRJNA1027780 | ENA |
REPOSITORIES: ENA
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Project description:Protection against pathogens is a major function of the gut microbiota. Although bacterial natural products have emerged as crucial components of host-microbiota interactions, their exact role in microbiota-mediated protection is largely unexplored. We addressed this knowledge gap with the nematodeCaenorhabditis elegansand its microbiota isolatePseudomonas fluorescensMYb115 that is known to protect againstBacillus thuringiensis (Bt) infection. We find that MYb115-mediated protection depends on sphingolipids that are derived from an iterative type I polyketide synthase (PKS), thereby describing a noncanonical pathway of bacterial sphingolipid production. We provide evidence that MYb115-derived sphingolipids affectC. eleganstolerance to Bt infection by altering host sphingolipid metabolism. This work establishes sphingolipids as structural outputs of bacterial PKS and highlights the role of microbiota-derived sphingolipids in host protection against pathogens.
2023-10-14 | GSE245296 | GEO
Project description:Knockout mutants of polyketide synthase genes were generated (pks1, pks2, pks3, pks14, pks18, pks37) and compared to wild-type AX2 strain of Dictyostelium discoideum using Agilent custom microarrays. The Dictyostelium cells were harvested at eight developmental stages (Amoeboid, Intiation of starvation, loose aggregate, Streaming, Mound, Slug, early culminant, Fruiting body) and subjected to microarray based expression profiling.
2015-07-18 | GSE71036 | GEO
Project description:Polyketide synthase function is a determinant of bacterial persistence and antibiotic tolerance in Burkholderia thailandensis
| PRJNA525635 | ENA
Project description:Persister cells are genetically identical variants in a bacterial population that have phenotypically modified their physiology to survive environmental stress. . To better define general persistence mechanisms that can be targeted to develop anti-persistence therapeutics, we performed transcriptomics analyses of Burkholderia thailandensis enriched for persisters using three methods: flow sorting by low proton motive force, meropenem treatment, and culture aging. Although the three persister populations generally expressed divergent gene expression profiles that underscores the multi-mechanistic nature of persistence, there were several common gene pathways expressed in two or more persister populations, including polyketide and non-ribosomal peptide synthesis, Clp proteases, mobile elements, enzymes involved in the lipid metabolism, and ATP-binding cassette (ABC) transporter systems. In particular, identification of genes that encode for polyketide synthases (PKS) and fatty acid catabolism indicate that generation of secondary metabolites, natural products, and complex lipids are needed for the maintenance of the persistence state. Persisters are markedly reduced in transposon mutants of the PKS gene BTH_I2366. Furthermore, treatment of multiple bacterial pathogens with the fatty acid synthesis inhibitor, CP-640186, potentiated antibiotic efficacy against the persisters. All together, our results suggest that bacterial persisters may exhibit an outward dormant physiology, but maintain active metabolic processes that are required to maintain persistence.
2019-03-06 | GSE127838 | GEO
Project description:Background: Among the nineteen polyketide synthases encoded by the genome of Mycobacterium tuberculosis, Pks13 was identified as the condensase required for the final condensation step of two long acyl chains. This leads to mycolic acids, which are essential components of the cell envelope of Corynebacterineae species. The single polypeptide chain of Pks13 comprises five catalytic domains, including two functional acyl carrier protein domains, separated by linkers comprising up to ca. 200 amino acid residues. In many aspects, Pks13 is an atypical polyketide synthase. It has been validated as a promising druggable target and its structure is badly needed to speed up drug discovery to fight against tuberculosis. Results: We report here a quasi-atomic model of Pks13 obtained using small angle X-ray scattering of the entire protein and various molecular subspecies combined with known high-resolution structures of Pks13 domains or structural homologues. In addition, the structural changes induced by the loading of a substrate analogue have been challenged. As a comparison, the low-resolution structures of two other mycobacterial polyketide synthases, Mas and PpsA from Mycobacterium bovis BCG, are also presented. This study highlights an identical and elongated structure of the apo and holo forms at the resolution probed. Catalytic domains are segregated into two parts, which correspond to the condensation reaction per se and to the release of the product, a pivot for the enzyme flexibility being at the interface. The two acyl carrier protein domains are found at opposite sides of the ketosynthase domain and display distinct characteristics in terms of flexibility. Apo-Pks13 is in our conditions in a monomeric state, but loading of a substrate analogue push the enzyme towards the functional dimeric state. Conclusions: The Pks13 model provides the first structural information on the molecular mechanism of this complex enzyme and open up new perspectives to develop inhibitors that target the interactions with its enzymatic partners or between catalytic domains within Pks13 itself.
2022-05-11 | PXD029754 | Pride
Project description:Mycobacterium polyketide synthase 13 (Pks13) is a protein complex comprised of a closely symmetric parallel dimer of chains, each encoding several enzymatic and transport functions. This module carries out the condensation of two different very long chain fatty acids to produce mycolic acids that are essential components of the mycobacterial cell wall. Here, we use cryo-EM, XL-MS, and integrative modeling to determine the structure and domain trajectories of the dimeric multi-enzyme Pks13 purified endogenously from mycobacteria under normal growth conditions. Structures of the multi-domain assembly revealed by cryogenic electron microscopy (cryoEM) define the ketosynthase (KS), linker, and acyltransferase (AT) domains, each at atomic resolution (1.8Å), with bound substrates defined at 2.4Å and 2.9Å resolution. Image classification reveals two distinct structures with alternate locations of the N-terminal acyl carrier protein (termed ACP1a, ACP1b) seen at 3.6Å and 4.6Å resolution respectively. These two structures suggest plausible intermediate states, related by a ~60Å movement of ACP1, on the pathway for substrate delivery from the fatty acyl-ACP ligase (FadD32) to the ketosynthase domain. The linking sequence between ACP1 and the KS includes an 11 amino acid sequence with 6 negatively charged side chains that lies in different positively charged grooves on the KS in ACP1a versus ACP1b structures. This charge complementarity between the extended chain and the grooves suggests some stabilization of these two distinct orientations. Other domains are visible at lower resolution and indicate flexibility relative to the KS-AT core. The chemical structures of three bound endogenous long chain fatty acid substrates with their proximal regions defined in the structures were determined by electrospray ionization mass spectrometry. The domain proximities were probed by chemical cross-linking and identified by mass spectrometry (XL-MS). These were incorporated into integrative structure modeling to define multiple domain configurations that transport the very long fatty acid chains throughout the multistep Pks13 mediated synthetic pathway.
2023-06-13 | PXD033471 | Pride
Project description:INC-Seq: Accurate single molecule reads using nanopore sequencing
| PRJEB12294 | ENA
Project description:Polyketide synthase (PKS) and nonribosomal peptide synthetase (NRPS) hybrid systems typically use complex protein-protein interactions to facilitate direct transfer of intermediates between these multimodular megaenzymes. In the canal-associated neurons (CANs) of Caenorhabditis elegans, PKS-1 and NRPS-1 produce the nemamides, the only known hybrid polyketide-nonribosomal peptides biosynthesized by animals, through a poorly understood mechanism. Here, we use genome editing and mass spectrometry to map the roles of individual PKS-1 and NRPS-1 enzymatic domains in nemamide biosynthesis. Furthermore, we show that nemamide biosynthesis requires at least five additional enzymes expressed in the CANs that are encoded by genes distributed across the worm genome. We identify the roles of these enzymes and discover a mechanism for trafficking intermediates between a PKS and an NRPS. Specifically, the enzyme PKAL-1 activates an advanced polyketide intermediate as an adenylate and directly loads it onto a carrier protein in NRPS-1. This trafficking mechanism provides a means by which a PKS-NRPS system can expand its biosynthetic potential and is likely important for the regulation of nemamide biosynthesis.
2021-06-22 | MTBLS2313 | MetaboLights
Project description:This data was acquired by engineering tri-,tetra- and pentaketide synthases using pikromycin synthase modules within K207-3 E. coli cells. The products were extracted with ethyl acetate, evaporate and re-suspended in 1:1 MeOH and H2O.
2024-05-31 | MSV000094913 | MassIVE