Traceless capping agent for peptide sequencing by partial edman degradation and mass spectrometry.
An improved method for the rapid sequence determination of biologically active peptides selected from one-bead-one-peptide combinatorial libraries has been developed. In this method, beads carrying unique peptide sequences were subjected to multiple cycles of partial Edman degradation (PED) by the treatment with a 15-30:1 mixture of phenyl isothiocyanate and N-(9-fluorenylmethoxycarbonyloxy)succinimide (Fmoc-OSU), to generate a series of sequence-specific truncation products (a peptide ladder) for each resin-bound peptide. Following PED, the Fmoc group was removed from the N-terminus and any reacted side chains by piperidine treatment. The sequence of the full-length peptide on each bead was then determined by matrix-assisted laser desorption ionization mass spectrometry. The use of Fmoc-OSU as a traceless capping agent resulted in cleaner MS spectra and improved reliability for sequence assignment. This rapid, sensitive, and inexpensive sequencing method should further expand the utility of combinatorial peptide libraries in biomedical research.
Thakkar
Amit
A
Department of Chemistry and Ohio State Biochemistry Program, Ohio State University, 100 West 18th Avenue, Columbus, Ohio 43210, USA.
Wavreille
Anne-Sophie
AS
Pei
Dehua
D
D000595:Amino Acid Sequence; D013058:Mass Spectrometry / Q000379:methods*; D008969:Molecular Sequence Data; D009943:Organophosphorus Compounds; D020539:Sequence Analysis, Protein / Q000379:methods*
C043517:2-(4-isothiocyanatophenoxy)-1,3,2-dioxaphosphinene 2-oxide; D009943:Organophosphorus Compounds