SynBio-SynChem Approaches to Diversifying the Pacidamycins through the Exploitation of an Observed Pictet-Spengler Reaction.

A nonenzymatic Pictet-Spengler reaction has been postulated to give rise to a subset of naturally occurring uridyl peptide antibiotics (UPAs). Here, using a combination of strain engineering and synthetic chemistry, we demonstrate that Pictet-Spengler chemistry may be employed to generate even greater diversity in the UPAs. We use an engineered strain to afford access to meta-tyrosine containing pacidamycin 4. Pictet-Spengler diversification of this compound using a small series of aryl-aldehydes was achieved with some derivatives affording remarkable diastereomeric control.

Caprazamycins: Biosynthesis and structure activity relationship studies.

Cell wall biosynthesis represents a valid target for antibacterial action but only a limited number of chemical structure classes selectively interact with specific enzymes or protein structures like transporters of the cell envelope. The integral membrane protein MraY translocase is essential for peptidoglycan biosynthesis catalysing the transfer of the peptidoglycan precursor phospho-MurNAc-pentapeptide to the lipid carrier undecaprenyl phosphate, thereby generating the cell wall intermediate lipid I. Not present in eukaryotic cells, MraY is a member of the superfamily of yet not well-understood integral membrane enzymes which involve proteins for bacterial lipopolysaccharide and teichoic acid or eukaryotic N-linked saccharides biosynthesis. Different natural nucleoside antibiotics as inhibitors of MraY translocase have been discovered comprising a glycosylated heterocyclic pyrimidin base among other potential lipid-, peptidic- or sugar moieties. Caprazamycins are liponucleoside antibiotics isolated from Streptomyces sp. MK730-62F2. They possess activity in vitro against Gram-positive bacteria, in particular against the genus Mycobacterium including M. intracellulare, M. avium and M. tuberculosis. Structural elucidation revealed the (+)-caprazol core skeleton as a unique moiety, the caprazamycins share with other MraY inhibitors such as the liposidomycins, A-90289 and the muraminomicins. They also share structural features such as uridyl-, aminoribosyl- and fatty acyl-moieties with other MraY translocase inhibitors like FR-900493 and the muraymycins. Intensive studies on their biosynthesis during the last decade identified not only common initial biosynthetic steps, but also revealed possible branching points towards individual biosynthesis of the respective compound. Structural diversity of caprazamycins was generated by feeding experiments, genetic engineering of the biosynthetic gene clusters and chemical synthesis for structure activity relationship studies with its target, MraY translocase.

The Systematic Investigation of the Quorum Sensing System of the Biocontrol Strain Pseudomonas chlororaphis subsp. aurantiaca PB-St2 Unveils aurI to Be a Biosynthetic Origin for 3-Oxo-Homoserine Lactones.

The shoot endophytic biocontrol strain Pseudomonas chlororaphis subsp. aurantiaca PB-St2 produces a wide range of exoproducts, including enzymes and antibiotics. The production of exoproducts is commonly tightly regulated. In order to get a deeper insight into the regulatory network of PB-St2, the strain was systematically investigated regarding its quorum sensing systems, both on the genetic and metabolic level. The genome analysis of PB-St2 revealed the presence of four putative acyl homoserine lactone (AHL) biosynthesis genes: phzI, csaI, aurI, and hdtS. LC-MS/MS analyses of the crude supernatant extracts demonstrated that PB-St2 produces eight AHLs. In addition, the concentration of all AHL derivatives was quantified time-resolved in parallel over a period of 42 h during the growth of P. aurantiaca PB-St2, resulting in production curves, which showed differences regarding the maximum levels of the AHLs (14.6 nM- 1.75 µM) and the production period. Cloning and heterologous overexpression of all identified AHL synthase genes in Escherichia coli proved the functionality of the resulting synthases PhzI, CsaI, and AurI. A clear AHL production pattern was assigned to each of these three AHL synthases, while the HdtS synthase did not lead to any AHL production. Furthermore, the heterologous expression study demonstrated unequivocally and for the first time that AurI directs the synthesis of two 3-oxo-AHLs.