title: Production of Sactipeptides in Escherichia coli: Probing the Substrate Promiscuity of Subtilosin A Biosynthesis.
abstract: Sactipeptides are peptide-derived natural products that are processed by remarkable, radical-mediated cysteine sulfur to α-carbon coupling reactions. The resulting sactionine thioether linkages give rise to the unique defined structures and concomitant biological activities of sactipeptides. An E. coli heterologous expression system, based on the biosynthesis of one such sactipeptide, subtilosin A, is described and this expression system is exploited to probe the promiscuity of the subtilosin A sactionine bond-forming enzyme, AlbA. These efforts allowed the facile expression and isolation of a small library of mutant sactipeptides based on the subtilosin A precursor peptide, demonstrating broad substrate promiscuity where none was previously known. Importantly, we show that the positions of the sactionine linkages can be moved, giving rise to new, unnatural sactipeptide structures. E. coli heterologous expression also allowed incorporation of unnatural amino acids into sactipeptides by means of amber-suppression technology, potentially opening up new chemistry and new applications for unnatural sactipeptides.
authors.affiliation: Division of Chemical Biology and Medicinal Chemistry, University of North Carolina at Chapel Hill, Eshelman School of Pharmacy , Chapel Hill, North Carolina, United States.
authors.forename: Paul M
authors.initials: PM
authors.lastname: Himes
authors.affiliation: Division of Chemical Biology and Medicinal Chemistry, University of North Carolina at Chapel Hill, Eshelman School of Pharmacy , Chapel Hill, North Carolina, United States.
authors.forename: Scott E
authors.initials: SE
authors.lastname: Allen
authors.affiliation: Division of Chemical Biology and Medicinal Chemistry, University of North Carolina at Chapel Hill, Eshelman School of Pharmacy , Chapel Hill, North Carolina, United States.
authors.forename: Sungwon
authors.initials: S
authors.lastname: Hwang
authors.affiliation: Division of Chemical Biology and Medicinal Chemistry, University of North Carolina at Chapel Hill, Eshelman School of Pharmacy , Chapel Hill, North Carolina, United States.
authors.forename: Albert A
authors.initials: AA
authors.lastname: Bowers
mesh_terms: D001430:Bacteriocins / Q000096:biosynthesis* / Q000235:genetics; D003545:Cysteine / Q000737:chemistry; D004926:Escherichia coli; D029968:Escherichia coli Proteins / Q000235:genetics / Q000378:metabolism; D015870:Gene Expression; D015394:Molecular Structure; D050862:Oxidoreductases Acting on Sulfur Group Donors / Q000235:genetics / Q000378:metabolism; D010456:Peptides, Cyclic / Q000096:biosynthesis* / Q000235:genetics; D015202:Protein Engineering; D011498:Protein Precursors / Q000096:biosynthesis / Q000235:genetics; D013379:Substrate Specificity
chemical_list: D001430:Bacteriocins; D029968:Escherichia coli Proteins; D010456:Peptides, Cyclic; D011498:Protein Precursors; C047828:sboA protein, Bacillus subtilis; C000622582:AlbA protein, E coli; D050862:Oxidoreductases Acting on Sulfur Group Donors; D003545:Cysteine