title: Synthesis of lucifensin by native chemical ligation and characteristics of its isomer having different disulfide bridge pattern.
abstract: The antimicrobial 40-amino-acid-peptide lucifensin was synthesized by native chemical ligation (NCL) using N-acylbenzimidazolinone (Nbz) as a linker group. NCL is a method in which a peptide bond between two discreet peptide chains is created. This method has been applied to the synthesis of long peptides and proteins when solid-phase synthesis is imcompatible. Two models of ligation were developed: [15+25] Ala-Cys and [19+21] His-Cys. The [19+21] His-Cys method gives lower yield because of the lower stability of 18-peptide-His-Nbz-CONH2 peptide, as suggested by density functional theory calculation. Acetamidomethyl-deprotection and subsequent oxidation of the ligated linear lucifensin gave a mixture of lucifensin isomers, which differed in the location of their disulfide bridges only. The dominant isomer showed unnatural pairing of cysteines [C1-6], [C3-5], and [C2-4], which limits its ability to form α-helical structure. The activity of isomeric lucifensin toward Bacillus subtilis, Staphylococcus aureus, and Micrococcus luteus was lower than that of the natural lucifensin. The desired product native lucifensin was prepared from this isomer using a one-pot reduction with dithiotreitol and subsequent air oxidation in slightly alkaline medium.
authors.affiliation: Institute of Organic Chemistry and Biochemistry, Czech Academy of Sciences, Flemingovo nam. 2, 16610, Prague, Czech Republic.
authors.forename: Stancho
authors.initials: S
authors.lastname: Stanchev
authors.forename: Zbigniew
authors.initials: Z
authors.lastname: Zawada
authors.forename: Lenka
authors.initials: L
authors.lastname: Monincová
authors.forename: Lucie
authors.initials: L
authors.lastname: Bednárová
authors.forename: Jiřina
authors.initials: J
authors.lastname: Slaninová
authors.forename: Vladimír
authors.initials: V
authors.lastname: Fučík
authors.forename: Václav
authors.initials: V
authors.lastname: Čeřovský
mesh_terms: D000890:Anti-Infective Agents / Q000138:chemical synthesis / Q000737:chemistry / Q000494:pharmacology; D023181:Antimicrobial Cationic Peptides / Q000138:chemical synthesis / Q000737:chemistry / Q000494:pharmacology; D023082:Defensins / Q000138:chemical synthesis / Q000737:chemistry / Q000494:pharmacology; D006094:Gram-Positive Bacteria / Q000254:growth & development*; D017433:Protein Structure, Secondary
chemical_list: D000890:Anti-Infective Agents; D023181:Antimicrobial Cationic Peptides; D023082:Defensins; C561217:lucifensin
keywords: CD spectroscopy; DTT reduction; antimicrobial activity; lucifensin; native chemical ligation; solid-phase peptide synthesis