ABSTRACT
Only a few natural products incorporating a diazeniumdiolate moiety have been isolated, and these compounds usually display a broad range of biological activities. Only recently has the first diazeniumdiolate natural product biosynthetic gene cluster been identified in Burkholderia cenocepacia H111, which produces the fungicide (-)-fragin and the signal molecule rac-valdiazen. In this study, l-valine was identified as the initial substrate of (-)-fragin biosynthesis with the aid of feeding experiments using isotopically labelled amino acid. The formation of the diazeniumdiolate was chemically studied with several proposed intermediates. Our results indicate that the functional group is formed during an early stage of the biosynthesis. Furthermore, an oxime compound was identified as a degradation product of (-)-fragin and was also observed in the crude extract of the wild-type strain. Moreover, a structure-activity relationship analysis revealed that each moiety of (-)-fragin is essential for its biological activity.
Subject(s)
Antifungal Agents/metabolism , Azo Compounds/metabolism , Bacterial Proteins/metabolism , Biological Products/metabolism , Burkholderia cenocepacia/enzymology , Genome, Bacterial , Oxidoreductases/metabolism , Antifungal Agents/chemistry , Antifungal Agents/pharmacology , Azo Compounds/chemistry , Azo Compounds/pharmacology , Bacterial Proteins/genetics , Biological Products/chemistry , Biological Products/pharmacology , Burkholderia cenocepacia/genetics , Gram-Negative Bacteria/drug effects , Gram-Negative Bacteria/growth & development , Gram-Positive Bacteria/drug effects , Gram-Positive Bacteria/growth & development , Microbial Sensitivity Tests , Multigene Family , Oxidoreductases/genetics , Structure-Activity Relationship , Substrate Specificity , Valine/chemistry , Valine/metabolismABSTRACT
Members of the diazeniumdiolate class of natural compounds show potential for drug development because of their antifungal, antibacterial, antiviral, and antitumor activities. Yet, their biosynthesis has remained elusive to date. Here, we identify a gene cluster directing the biosynthesis of the diazeniumdiolate compound fragin in Burkholderia cenocepacia H111. We provide evidence that fragin is a metallophore and that metal chelation is the molecular basis of its antifungal activity. A subset of the fragin biosynthetic genes is involved in the synthesis of a previously undescribed cell-to-cell signal molecule, valdiazen. RNA-Seq analyses reveal that valdiazen controls fragin biosynthesis and affects the expression of more than 100 genes. Homologs of the valdiazen biosynthesis genes are found in various bacteria, suggesting that valdiazen-like compounds may constitute a new class of signal molecules. We use structural information, in silico prediction of enzymatic functions and biochemical data to propose a biosynthesis route for fragin and valdiazen.
Subject(s)
Azo Compounds/metabolism , Burkholderia cenocepacia/metabolism , Antifungal Agents/metabolism , Antifungal Agents/pharmacology , Azo Compounds/pharmacology , Burkholderia cenocepacia/genetics , Chelating Agents/metabolism , Chelating Agents/pharmacology , Homeostasis , Multigene Family , Quorum SensingABSTRACT
Synthetic studies toward highly oxygenated seco-prezizaane sesquiterpenes are reported, which culminated in a formal total synthesis of the neurotrophic agent (-)-jiadifenolide. For the construction of the tricyclic core structure, an unusual intramolecular and diastereoselective Nozaki-Hiyama-Kishi reaction involving a ketone as electrophilic coupling partner was developed. In addition, synthetic approaches toward the related natural product (2R)-hydroxy-norneomajucin, featuring a Mn-mediated radical cyclization for the tricycle assembly and a regioselective OH-directed C-H activation are presented.
Subject(s)
Sesquiterpenes/chemistry , Carbon-13 Magnetic Resonance Spectroscopy , Cyclization , Proton Magnetic Resonance Spectroscopy , Sesquiterpenes/chemical synthesis , Spectrometry, Mass, Electrospray Ionization , StereoisomerismABSTRACT
A formal [2 + 2 + 2] cycloaddition reaction between a 1,3-dione, an olefin, and molecular oxygen mediated by light is reported, which delivers endoperoxides in good yield through the formation of two C-O and one C-C bond in one step. The resulting 1,2-dioxanes are stable compounds and can be further derivatized at the hemiacetal position via alkylation or acetylation. All compounds have been evaluated against Plasmodium falciparum, and the best compound displayed an IC50-value of 180 nM. A potential mechanistic rationale for the formation of these compounds is presented.
