Biology subjects

Schalk, I. J.

Publications and source records attributed to Schalk, I. J..

5 recordsLinked to original sources

bioRxiv2026

PqqU (PA2289) is responsible for Pyrroloquinoline Quinone Uptake in Pseudomonas aeruginosa

Pseudomonas aeruginosa relies on the redox cofactor pyrroloquinoline quinone (PQQ) for efficient glucose and ethanol metabolism via periplasmic dehydrogenases (Gcd and ExaA). While PQQ biosynthesis is well-characterized, its uptake mechanisms remain unclear. Here, we identify PA2289 (PqqU), a TonB-dependent transporter, as the primary PQQ importer in P. aeruginosa. Growth assays with PQQ-deficient mutants ({Delta}pqqABCDEH) demonstrated that PqqU is essential for exogenous PQQ uptake, rescuing growth on glucose and ethanol. Genomic analysis across 210 P. aeruginosa and 263 Pseudomonas strains revealed high conservation of PQQ biosynthesis and utilization genes, while PqqU showed lower prevalence (47.7%) in the genus. Transcriptional analyses using fluorescent reporters and qRT-PCR demonstrated that PqqU expression remains unchanged in response to PQQ, varying carbon sources, or iron availability, suggesting constitutive regulation. Comparative proteomics between wild-type and {Delta}pqqABCDEH strains, cultured on glucose or ethanol, uncovered extensive proteomic shifts, underscoring P. aeruginosas metabolic adaptability. Additionally, PQQ-dependent metabolic pathways appear to indirectly influence iron homeostasis, most likely through environmental acidification. Together, these results emphasize the critical role of PqqU in PQQ uptake and its broader significance in shaping the metabolic and environmental versatility of Pseudomonas.

microbiology↗

bioRxiv2025

Salmonella relies on siderophore exploitation at low pH

Salmonella enterica, a prominent enteric pathogen, employs sophisticated iron acquisition mechanisms to overcome host-imposed iron limitation, notably through the production and uptake of siderophores--small, high-affinity iron-chelating compounds that scavenge iron from the host environment. In this study, we investigate how environmental pH influences Salmonellas preference for its endogenous siderophores versus exogenous siderophores within the physiological range of the gastrointestinal tract. Through competition assays, gene expression analysis, and siderophore quantification, we demonstrate that Salmonella increasingly relies on exogenous siderophores under acidic conditions. This shift is attributed to reduced production of its endogenous siderophores, enterobactin and salmochelin. Deletion of the sigma factor RpoS enhances endogenous siderophore production and iron acquisition at low pH, suggesting its role in regulating iron homeostasis. Our findings reveal a pH-dependent adaptation in Salmonellas iron acquisition strategy, highlighting the pathogens versatility in nutrient acquisition across varying gastrointestinal conditions. This research provides insights into Salmonellas pathogenicity and may inform the development of targeted interventions for Salmonella infections.

microbiology↗

bioRxiv2024

Characterisation of Pseudomonas aeruginosas metal-responsive TonB-dependent transporters

Pseudomonas aeruginosa, a versatile bacterium, relies on several TonB-dependent transporters (TBDTs) for nutrient acquisition (such as iron-siderophore complexes) and adaptation to various environments. While some TBDTs are well characterized, a significant number remain unexplored despite their potential role in pathogenicity. In this study, we developed fluorescent reporter plasmids to investigate TBDT promoter activity. Initially, we characterized their promoter activity in commonly used laboratory conditions, revealing diverse expression patterns among all TBDTs. Subsequently, we classified the TBDTs into distinct metal-responsive groups based on their stress-responsive behaviour, shedding light on their functional roles. Additionally, we show that these reporter constructs can be used as a powerful tool for siderophore detection. Finally, single cell analysis of TBDT promoter activity during coculture with an enterobactin-producing Klebsiella pneumoniae strain shows homogenous expression of key TBDT in P. aeruginosa. Our findings provide valuable insights into the expression profiles and functional diversity of P. aeruginosa TBDTs in complex conditions, reaching from commonly used lab media to complex co-culturing conditions.

microbiology↗

bioRxiv2023

PvdL Orchestrates the Assembly of the Nonribosomal Peptide Synthetases Involved in Pyoverdine Biosynthesis in Pseudomonas aeruginosa.

The pyoverdine siderophore is produced by Pseudomonas aeruginosa to access iron. Its synthesis involves the complex co-ordination of four nonribosomal peptide synthetases (NRPSs), responsible for assembling the pyoverdine peptide backbone. The precise cellular organization of these NRPSs and their mechanisms of interaction remain unclear. Here, we used a combination of several single-molecule microscopy techniques to to elucidate the spatial arrangement of NRPSs within pyoverdine-producing cells. Our findings revealed that PvdL differs from the three other NRPS in term of localization and mobility patterns. PvdL is predominantly located at the inner membrane, while the other also explore the cytoplasmic compartment. Leveraging the power of multicolor single-molecule localization, we further reveal co-localization between PvdL and the other NRPSs, suggesting a pivotal role for PvdL in orchestrating the intricate biosynthetic pathway. Our observations strongly indicates that PvdL serves as a central orchestrator in the assembly of NRPSs involved in pyoverdine biosynthesis, assuming a critical regulatory function.

microbiology↗

bioRxiv2018

Macromolecular organization of the pyoverdine biosynthetic pathway in Pseudomonas aeruginosa.

The biosynthesis of the pyoverdine siderophore in Pseudomonas aeruginosa inlvolves multiple enzymes including Non-Ribosomal Peptide Synthetases (NRPS). We used in vitro single-molecule tracking and FRET-FLIM (Forster resonance energy transfer measured by fluorescence lifetime microscopy) to explore the spatial partitioning of the ornithine hydroxylase PvdA and to characterize its interactions with NRPS. The statistical description of thousands of single PvdA traces in cells using jump distance distribution analysis showed PvdA was mostly diffusing bound to large complex in the cytoplasm with a small exchangeable trapped fraction diffusing slower. FRET-FLIM clearly showed PvdA are physically interacting with the four NRPS PvdJ, PvdI, PvdL and PvdD of the pyoverdine pathway in the cellular context. Our data provide evidence for strongly organized multi-enzymatic biosynthetic complexes named siderosomes responsible for the siderophore biosynthesis. PvdA binding mode is strikingly different according to the NRPS it is interacting with suggesting PvdA binding site have co-evolved with the enzymatic active sites of NRPS.

biophysics↗