Characterization of Sviceucin from Streptomyces Provides Insight into Enzyme Exchangeability and Disulfide Bond Formation in Lasso Peptides.
Lasso peptides are bacterial ribosomally synthesized and post-translationally modified peptides. They have sparked increasing interest in peptide-based drug development because of their compact, interlocked structure, which offers superior stability and protein-binding capacity. Disulfide bond-containing lasso peptides are rare and exhibit highly sought-after activities. In an effort to expand the repertoire of such molecules, we heterologously expressed, in Streptomyces coelicolor, the gene cluster encoding sviceucin, a type I lasso peptide with two disulfide bridges originating from Streptomyces sviceus, which allowed it to be fully characterized. Sviceucin and its reduced forms were characterized by mass spectrometry and peptidase digestion. The three-dimensional structure of sviceucin was determined using NMR. Sviceucin displayed antimicrobial activity selectively against Gram-positive bacteria and inhibition of fsr quorum sensing in Enterococcus faecalis. This study adds sviceucin to the type I lasso peptide family as a new representative. Moreover, new clusters encoding disulfide-bond containing lasso peptides from Actinobacteria were identified by genome mining. Genetic and functional analyses revealed that the formation of disulfide bonds in sviceucin does not require a pathway-encoded thiol-disulfide oxidoreductase. Most importantly, we demonstrated the functional exchangeability of the sviceucin and microcin J25 (a non-disulfide-bridged lasso peptide) macrolactam synthetases in vitro, highlighting the potential of hybrid lasso synthetases in lasso peptide engineering.
Laboratory Molecules of Communication and Adaptation of Microorganisms (MCAM, UMR 7245 CNRS-MNHN), Sorbonne Universités, Muséum National d'Histoire Naturelle, Centre National de la Recherche Scientifique, CP 54, 57 rue Cuvier, F-75005, Paris, France.
Yanyan
Y
Li
Laboratory Molecules of Communication and Adaptation of Microorganisms (MCAM, UMR 7245 CNRS-MNHN), Sorbonne Universités, Muséum National d'Histoire Naturelle, Centre National de la Recherche Scientifique, CP 54, 57 rue Cuvier, F-75005, Paris, France.
Rémi
R
Ducasse
Laboratory Molecules of Communication and Adaptation of Microorganisms (MCAM, UMR 7245 CNRS-MNHN), Sorbonne Universités, Muséum National d'Histoire Naturelle, Centre National de la Recherche Scientifique, CP 54, 57 rue Cuvier, F-75005, Paris, France.
Séverine
S
Zirah
Laboratory Molecules of Communication and Adaptation of Microorganisms (MCAM, UMR 7245 CNRS-MNHN), Sorbonne Universités, Muséum National d'Histoire Naturelle, Centre National de la Recherche Scientifique, CP 54, 57 rue Cuvier, F-75005, Paris, France.
Alain
A
Blond
Laboratory Molecules of Communication and Adaptation of Microorganisms (MCAM, UMR 7245 CNRS-MNHN), Sorbonne Universités, Muséum National d'Histoire Naturelle, Centre National de la Recherche Scientifique, CP 54, 57 rue Cuvier, F-75005, Paris, France.
Christophe
C
Goulard
Institut de Chimie des Substances Naturelles, Centre de Recherche de Gif, UPR 2301 CNRS Université Paris-Sud, 1 avenue de la Terrasse, F-91198 Gif-sur-Yvette, France.
Ewen
E
Lescop
Unité de Recherche Risques Microbiens (U2RM)-Stress et Virulence (EA 4655), Université de Caen-Basse Normandie, F-14032 Caen, France.
Caroline
C
Giraud
Unité de Recherche Risques Microbiens (U2RM)-Stress et Virulence (EA 4655), Université de Caen-Basse Normandie, F-14032 Caen, France.
Axel
A
Hartke
Institut de Chimie des Substances Naturelles, Centre de Recherche de Gif, UPR 2301 CNRS Université Paris-Sud, 1 avenue de la Terrasse, F-91198 Gif-sur-Yvette, France.
Eric
E
Guittet
Institute for Integrative Biology of the Cell (I2BC), CEA, CNRS, Université Paris-Sud, Bât. 400, Université Paris-Sud, F-91405 Orsay, France.
Jean-Luc
JL
Pernodet
Laboratory Molecules of Communication and Adaptation of Microorganisms (MCAM, UMR 7245 CNRS-MNHN), Sorbonne Universités, Muséum National d'Histoire Naturelle, Centre National de la Recherche Scientifique, CP 54, 57 rue Cuvier, F-75005, Paris, France.
Sylvie
S
Rebuffat
D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000737:chemistry* / Q000235:genetics* / Q000378:metabolism; D002851:Chromatography, High Pressure Liquid; D004220:Disulfides / Q000737:chemistry; D009682:Magnetic Resonance Spectroscopy; D008958:Models, Molecular; D008969:Molecular Sequence Data; D005810:Multigene Family; D010455:Peptides / Q000737:chemistry / Q000378:metabolism*; D016415:Sequence Alignment; D013302:Streptomyces / Q000737:chemistry* / Q000201:enzymology / Q000235:genetics
D001426:Bacterial Proteins; D004220:Disulfides; D010455:Peptides