Recombineering enables genome mining of novel siderophores in a non-model Burkholderiales strain

Iron is essential for bacterial survival,and most bacteria capture iron by producing siderophores.Burkholde-riales bacteria produce various types of bioactive secondary metabolites,such as ornibactin and malleobactin siderophores.In this study,the genome analysis of Burkholderiales genomes showed a putative novel siderophore gene cluster crb,which is highly similar to the ornibactin and malleobactin gene clusters but does not have pvdF,a gene encoding a formyltransferase for N-δ-hydroxy-ornithine formylation.Establishing the bacteriophage recom-binase Redγ-Redδβ7029 mediated genome editing system in a non-model Burkholderiales strain Paraburkholderia caribensis CICC 10960 allowed the rapid identification of the products of crb gene cluster,caribactins A-F(1-6).Caribactins contain a special amino acid residue N-δ-hydroxy-N-δ-acetylornithine(haOrn),which differs from the counterpart N-δ-hydroxy-N-δ-formylornithine(hOrn)in ornibactin and malleobactin,owing to the absence of pvdF.Gene inactivation showed that the acetylation of hOrn is catalyzed by CrbK,whose homologs proba-bly not be involved in the biosynthesis of ornibactin and malleobactin,showing possible evolutionary clues of these siderophore biosynthetic pathways from different genera.Caribactins promote biofilm production and en-hance swarming and swimming abilities,suggesting that they may play crucial roles in biofilm formation.This study also revealed that recombineering has the capability to mine novel secondary metabolites from non-model Burkholderiales species.

Development of environmentally friendly biological algicide and biochemical analysis of inhibitory effect of diatom Skeletonema costatum

Skeletonema costatum is a diatom widely distributed in red tide microalgae blooms and as one of the main algae causing harmful algal blooms,because of their rapid reproduction and production of toxic and harmful substances,often play a negative role in aquatic ecosystems,and human health and wellbeing.Bacillomycin D is a nonribosomal cyclic antifungal lipopeptide in the iturins family.In this study,Bacillomycin D was tested for its ability to inhibit the growth of S.costatum.The EC_(50 24h) of Bacillomycin D on S.costatum was 24.70μg/mL.The chlorophyll fluorescence parameters F_(v)/F_(m),F_(v)/F_(o),and yield of the diatoms decreased significantly with increasing concentrations of Bacillomycin D.Study of the mechanism showed that Bacillomycin D induced cell death by changing cell membrane permeability,promoting the release of cellular contents.In this study,transcriptomic analysis showed Bacillomycin D significantly inhibited the photosynthesis and metabolism of S.costatum.These findings investigated the inhibitory effect of Bacillomycin D on the growth of S.costatum and provided a theoretical foundation for the development of new environmentally friendly biological algicide.

In vitro characterization of a nitro-forming oxygenase involved in 3-(trans-2’-aminocyclopropyl)alanine biosynthesis

In vitro characterization experiments revealed the formations of 3-(trans-2’-aminocyclopropyl)alanine((3-Acp)Ala)and 3-(trans-2’-nitrocyclopropyl)alanine((3-Ncp)Ala)are originated via two homologous proteins,BelK and HrmI,which regioselectively catalyze the N𝜀-oxygenation of l-lysine.The two enzymes belong to the emerg-ing heme-oxygenase-like diiron oxidase and oxygenase(HDO)superfamily and the catalytic center of BelK is validated by homology modeling and site-directed mutations.Based on the in vitro characterization,the biosyn-thetic pathways of(3-Acp)Ala and(3-Ncp)Ala are proposed.