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Molecular mechanisms of nutrient uptake in marine microbial ecosystems
thesisposted on 2022-03-29, 03:15 authored by Andrew George McLeish
Unicellular cyanobacteria are highly diverse, abundant and ubiquitous in the global oceans. Investigating the range and magnitude of organic compound transport in these organisms will help define pathways of energy and nutrient cycling in marine ecosystems.ABC membrane transport systems utilise a periplasmic binding protein (PBP) to confer substrate specificity. These PBPs are thus ideal targets to study the ecological significance of predicted amino acid transporters. Twelve PBPs were investigated that group into three orthologous gene clusters with putative ligands: branched-chain amino acids;carbohydrates and amino acids; or amino acids, based on homology predictions.A high-throughput recombinant expression strategy enabled a protein from each cluster to be successfully heterologously expressed and purified. Differential scanning fluorimetry (DSF) analysis revealed that "SW840" binds a broad range of amino acids, aspredicted. "SC634" on the other hand was found to bind with formate and formaldehyde, not the predicted ligand of branched-chain amino acids.This work provides important evidence of the specific ligand binding characteristics for each transporter. Future work to elucidate the cellular fate of these compounds is proposed. The ability of cyanobacteria, often considered to be obligatory phototrophs, tosalvage organic compounds may provide a competitive advantage in marine ecosystems.