title: Efficient conjugation of peptides to oligonucleotides by "native ligation".
abstract: A new strategy has been developed for conjugation of peptides to oligonucleotides. The method is based on the "native ligation" of an N-terminal thioester-functionalized peptide to a 5'-cysteinyl oligonucleotide. Two new reagents were synthesized for use in solid-phase peptide and oligonucleotide synthesis, respectively. Pentafluorophenyl S-benzylthiosuccinate was used in the final coupling step in standard Fmoc-based solid-phase peptide assembly. Deprotection with trifluoracetic acid generated in solution peptides substituted with an N-terminal S-benzylthiosuccinyl moiety. O-trans-4-(N-alpha-Fmoc-S-tert-butylsulfenyl-L-cysteinyl)aminoc yclohe xyl O-2-cyanoethyl-N,N-diisopropylphosphoramidite was used in the final coupling step in standard phosphoramidite solid-phase oligonucleotide assembly. Deprotection with aqueous ammonia solution generated in solution 5'-S-tert-butylsulfenyl-L-cysteinyl functionalized oligonucleotides. Functionalized peptides and oligonucleotides were used without purification in native ligation conjugation reactions in aqueous/organic solution using tris-(2-carboxyethyl)phosphine to remove the tert-butylsulfenyl group in situ and thiophenol as a conjugation enhancer. A range of peptide-oligonucleotide conjugates were prepared by this route and purified by reversed-phase HPLC.
authors.affiliation: Medical Research Council, Laboratory of Molecular Biology, Hills Road, Cambridge, CB2 2QH, U.K.
authors.forename: D A
authors.initials: DA
authors.lastname: Stetsenko
authors.forename: M J
authors.initials: MJ
authors.lastname: Gait
mesh_terms: D000595:Amino Acid Sequence; D002851:Chromatography, High Pressure Liquid; D008956:Models, Chemical; D008969:Molecular Sequence Data; D009841:Oligonucleotides / Q000378:metabolism*; D010455:Peptides / Q000138:chemical synthesis / Q000378:metabolism*; D014269:Trifluoroacetic Acid
chemical_list: D009841:Oligonucleotides; D010455:Peptides; D014269:Trifluoroacetic Acid