RESUMO
Selenium is an essential micronutrient in diverse organisms. Two routes are known for its insertion into proteins and nucleic acids, via selenocysteine and 2-selenouridine, respectively1. However, despite its importance, pathways for specific incorporation of selenium into small molecules have remained elusive. Here we use a genome-mining strategy in various microorganisms to uncover a widespread three-gene cluster that encodes a dedicated pathway for producing selenoneine, the selenium analogue of the multifunctional molecule ergothioneine2,3. We elucidate the reactions of all three proteins and uncover two novel selenium-carbon bond-forming enzymes and the biosynthetic pathway for production of a selenosugar, which is an unexpected intermediate en route to the final product. Our findings expand the scope of biological selenium utilization, suggest that the selenometabolome is more diverse than previously thought, and set the stage for the discovery of other selenium-containing natural products.
Assuntos
Vias Biossintéticas , Genes Microbianos , Histidina/análogos & derivados , Compostos Organosselênicos , Selênio , Produtos Biológicos/química , Produtos Biológicos/metabolismo , Vias Biossintéticas/genética , Carbono/metabolismo , Enzimas , Ergotioneína , Genes Microbianos/genética , Histidina/biossíntese , Metaboloma/genética , Micronutrientes/biossíntese , Família Multigênica/genética , Proteínas , Selênio/metabolismoRESUMO
Selenium (Se) is an essential micronutrient that is found naturally in proteins, nucleic acids, and natural products. Unlike selenoproteins and selenonucleic acids, little is known about the structures of biosynthetic enzymes that incorporate Se into small molecules. Here, we report the X-ray crystal structure of SenB, the first known Se-glycosyltransferase that was recently found to be involved in the biosynthesis of the Se-containing metabolite selenoneine. SenB catalyzes C-Se bond formation using selenophosphate and an activated uridine diphosphate sugar as a Se and glycosyl donor, respectively, making it the first known selenosugar synthase and one of only four bona fide C-Se bond-forming enzymes discovered to date. Our crystal structure, determined to 2.25 Å resolution, reveals that SenB is a type B glycosyltransferase, displaying the prototypical fold with two globular Rossmann-like domains and a catalytic interdomain cleft. By employing complementary structural biology techniques, we find that SenB undergoes both local and global substrate-induced conformational changes, demonstrating a significant increase in α-helicity and a transition to a more compact conformation. Our results provide the first structure of SenB and set the stage for further biochemical characterization in the future.
Assuntos
Selênio , Selênio/metabolismo , Glicosiltransferases , Ligantes , Selenoproteínas , Cristalografia por Raios XRESUMO
Phosphonate natural products are renowned for inhibitory activities which underly their development as antibiotics and pesticides. Although most phosphonate natural products have been isolated from Streptomyces, bioinformatic surveys suggest that many other bacterial genera are replete with similar biosynthetic potential. While mining actinobacterial genomes, we encountered a contaminated Mycobacteroides data set which included a biosynthetic gene cluster predicted to produce novel phosphonate compounds. Sequence deconvolution revealed that the contig containing this cluster, as well as many others, belonged to a contaminating Bacillus and is broadly conserved among multiple species, including the epiphyte Bacillus velezensis. Isolation and structure elucidation revealed a new di- and tripeptide composed of l-alanine and a C-terminal l-phosphonoalanine which we name phosphonoalamides E and F. These compounds exhibit broad-spectrum antibacterial activity, including strong inhibition against the agricultural pests responsible for vegetable soft rot (Erwinia rhapontici), onion rot (Pantoea ananatis), and American foulbrood (Paenibacillus larvae). This work expands our knowledge of phosphonate metabolism and underscores the importance of including underexplored microbial taxa in natural product discovery. IMPORTANCE Phosphonate natural products produced by bacteria have been a rich source of clinical antibiotics and commercial pesticides. Here, we describe the discovery of two new phosphonopeptides produced by B. velezensis with antibacterial activity against human and plant pathogens, including those responsible for widespread soft rot in crops and American foulbrood. Our results provide new insight on the natural chemical diversity of phosphonates and suggest that these compounds could be developed as effective antibiotics for use in medicine or agriculture.
Assuntos
Anti-Infecciosos , Bacillus , Produtos Biológicos , Organofosfonatos , Praguicidas , Humanos , Produtos Biológicos/química , Bacillus/genética , Bacillus/metabolismo , Antibacterianos/farmacologia , Antibacterianos/química , Bactérias/genética , Genoma BacterianoRESUMO
The trace element selenium (Se) is essential to the physiology of most organisms on the planet. The most well documented of Se's biological forms are selenoproteins, where selenocysteine often serves as the catalytic center for crucial redox processes. Se is also found in several other classes of biological molecules, including nucleic acids, sugars, and modified amino acids, although its role in the function of these metabolites is less understood. Despite its prevalence, only a small number of Se-specific biosynthetic pathways have been discovered. Around half of these were first characterized in the past three years, suggesting that the selenometabolome may be more diverse than previously appreciated. Here, we review the recent advances in our understanding of this intriguing biochemical space, and discuss prospects for future discovery efforts.
Assuntos
Selênio , Selenoproteínas , Selênio/metabolismo , Selênio/química , Selenoproteínas/metabolismo , Humanos , Animais , Selenocisteína/metabolismo , Selenocisteína/química , Enzimas/metabolismo , Oxirredução , Ácidos Nucleicos/metabolismo , Ácidos Nucleicos/químicaRESUMO
Selenium is an essential micronutrient, but its presence in biology has been limited to protein and nucleic acid biopolymers. The recent identification of a biosynthetic pathway for selenium-containing small molecules suggests that there is a larger family of selenometabolites that remains to be discovered. Here we identify a recently evolved branch of abundant and uncharacterized metalloenzymes that we predict are involved in selenometabolite biosynthesis using a bioinformatic search strategy that relies on the mapping of composite active site motifs. Biochemical studies confirm this prediction and show that these enzymes form an unusual C-Se bond onto histidine, thus giving rise to a distinct selenometabolite and potent antioxidant that we have termed ovoselenol. Aside from providing insights into the evolution of this enzyme class and the structural basis of C-Se bond formation, our work offers a blueprint for charting the microbial selenometabolome in the future.
RESUMO
Selenium is an essential micronutrient, but its presence in biology has been limited to protein and nucleic acid biopolymers. The recent identification of the first biosynthetic pathway for selenium-containing small molecules suggests that there is a larger family of selenometabolites that remains to be discovered. Using a bioinformatic search strategy that relies on mapping of composite active site motifs, we identify a recently evolved branch of abundant and uncharacterized metalloenzymes that we predict are involved in selenometabolite biosynthesis. Biochemical studies confirm this prediction and show that these enzymes form an unusual C-Se bond onto histidine, thus giving rise to a novel selenometabolite and potent antioxidant that we have termed ovoselenol. Aside from providing insights into the evolution of this enzyme class and the structural basis of C-Se bond formation, our work offers a blueprint for charting the microbial selenometabolome in the future.
RESUMO
Non-heme iron-dependent sulfoxide/selenoxide synthases (NHISS) constitute a unique metalloenzyme class capable of installing a C-S/Se bond onto histidine to generate thio/selenoimidazole antioxidants, such as ergothioneine and ovothiol. These natural products are increasingly recognized for their health benefits. Among associated ergothioneine-biosynthetic enzymes, type IV EgtBs stand out, as they exhibit low sequence similarity with other EgtB subfamilies due to their recent divergence from the ovothiol-biosynthetic enzyme OvoA. Herein, we present crystal structures of two representative EgtB-IV enzymes, offering insights into the basis for this evolutionary convergence and enhancing our understanding of NHISS active site organization more broadly. The ability to interpret how key residues modulate substrate specificity and regioselectivity has implications for downstream identification of divergent reactivity within the NHISS family. To this end, we identify a previously unclassified clade of OvoA-like enzymes with a seemingly hybrid set of characteristics, suggesting they may represent an evolutionary intermediate between OvoA and EgtB-IV.
RESUMO
Phosphonic acid natural products have potent inhibitory activities that have led to their application as antibiotics. Recent studies uncovered large collections of gene clusters encoding for unknown phosphonic acids across microbial genomes. However, our limited understanding of their metabolism presents a significant challenge toward accurately informing the discovery of new bioactive compounds directly from sequence information alone. Here, we use genome mining to identify a family of gene clusters encoding a conserved branch point unknown to bacterial phosphonic acid biosynthesis. The products of this gene cluster family are the phosphonoalamides, four new phosphonopeptides with l-phosphonoalanine as the common headgroup. Phosphonoalanine and phosphonoalamide A are antibacterials, with strongest inhibition observed against strains of Bacillus and Escherichia coli. Heterologous expression identified the gene required for transamination of phosphonopyruvate to phosphonoalanine, a new route for bacterial phosphonic acids encoded within genomes of diverse microbes. These results expand our knowledge of phosphonic acid diversity and pathways for their biosynthesis.