Chemical and Genomic Analysis of Siderophores of Pseudomonas fluorescens YK-310

Abstract

Siderophores are fascinating secondary metabolites produced by bacteria primarily under iron-deficient conditions to obtain the vital trace element iron. However, their significance extends far beyond this primary function. In agriculture, for example, they exert a remarkable effect against phytopathogens, while in pharmaceutics they find application both in the treatment of heavy metal poisoning and as promising antibiotic conjugates. Pyoverdines belong to the siderophores, of which more than 60 different structures have been elucidated to date, although the structures of many representatives of this compound class remain unsolved. In the context of this doctoral thesis, the pyoverdine of the strain Pseudomonas fluorescens YK-310 was therefore characterised structurally. For this purpose, the associated gene cluster was investigated using bioinformatic tools, and analytical methods such as high-performance liquid chromatography (HPLC), liquid chromatography coupled with mass spectrometry (LC-MS), and nuclear magnetic resonance spectroscopy (NMR) were employed. The successful structure elucidation yielded a pyoverdine which can thus be added as a further representative to the growing collection of known siderophores.

Another topic of this doctoral thesis is ferrocin, a natural product likewise produced by the strain Pseudomonas fluorescens YK-310, whose possible role as a siderophore was investigated, among other things, within the scope of this work. Initially, the focus was on the downstream processes of the compound, which were carried out by means of HPLC to obtain sufficient pure substance for the studies on the mechanism of action and the modifications. The mode of action underlying the antibiotic activity of ferrocin has previously remained unclear and was therefore to be investigated in greater detail. In addition, the influence of fatty acid side chains of varying lengths on its efficacy was unknown, which prompted an investigation into whether ferrocin can be chemically and enzymatically deacylated and how different fatty acid chains affect its biological activity. In addition to HPLC and LC-MS, biological activity assays were conducted for this purpose. The investigations revealed that ferrocin interferes with lipoprotein biosynthesis, although additional mechanisms of action cannot be ruled out. The chemical deacylation of ferrocin was accomplished successfully, enabling the establishment of a corresponding methodology. Enzymatic deacylation of the intact form, however, was unsuccessful. Finally, it could be demonstrated that ferrocin is not produced under iron-deficient conditions, environments under which siderophores are normally produced in greater quantities, and therefore ferrocin cannot be classified among the classical siderophores.

Die Dissertation ist gesperrt bis zum 22. Juli 2028 !