A study on amphiphilic siderophore detection, structure elucidation and their iron-mediated vesicle self-assembly
Serrano Figueroa, Luis O'mar.
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Soap Lake, located in Washington State, was the subject of an NSF funded Microbial Observatory and is a naturally occurring saline and alkaline lake. Several organisms inhabiting this lake have been identified as producers of siderophores that are unique in structure. Two isolates SL01 & SL28 were the focus of this study of siderophore production, structure elucidation and vesicle self-assembly. Bacterial isolates, enriched from Soap Lake sediment and water samples, were screened for siderophore production. Siderophore production was confirmed through the chrome azurol S (CAS) agar plate method. Isolates SL01 and SL28 were found to produce relatively high concentrations of siderophores in liquid medium. Extraction was performed by the methanol/water protocol in Varian cartridges and siderophore purification was done on HPLC with a 0-70% acetonitrile gradient. Lyophilization or in vacuo evaporation followed in order to store siderophores. Siderophore structure was determined using liquid chromatography and tandem mass spectrometry (LC/MS/MS) with fatty acid methyl ester (FAME) analysis. Vesicle self-assembly studies were performed using dynamic light scattering (DLS) and epifluorescence microscopy (employing cryoembedding and cryosectioning). Three new amphiphilic siderophore families (two from SL01 and one from SL28) were produced by the bacterial isolates, found to be most closely related to Halomonas variablis and Halomonas pantelleriensis, respectively. These siderophores resemble the amphiphilic aquachelin siderophores produced by Halomonas aquamarina strain DS40M3, a marine bacterium. Addition of ferric iron (Fe +3) at different equivalents demonstrated vesicle formation and this was confirmed by both DLS and epifluorescence microscopy. Bacteria thriving under saline and alkaline conditions are capable of producing unique siderophores resembling those produced by microbes inhabiting marine environments. Vesicle self-assembly was confirmed quantitatively and qualitatively. Amphiphilic siderophores may have different applications in medical and environmental fields.