DEVELOPMENT OF AN EFFICIENT AMINO-ACID SEQUENCING METHOD USING FLUORESCENT EDMAN REAGENT 7-[(N,N-DIMETHYLAMINO)SULFONYL]-2,1,3-BENZOXADIAZOL-4-YL ISOTHIOCYANATE

In this paper, a new method is described for N-terminal amino acid sequencing of peptides using the fluorescent reagent 7-[(N,N-dimethylamino)sulfonyl]-2,1,3-benzoxadiazol-4-yl isothiocyanate (DBD-NCS). Sequence determination is carried out by identifying thiazolinone (TZ) amino acids, which are generally unstable and difficult to detect. The employed system can easily and quickly derive TZ amino acids using the Edman reaction with DBD-NCS; these amino acids are also stable enough to be efficiently detected by high-performance liquid chromatography. Resultant detection limits for DBD-TZ amino acids range from 50 fmol to a sub-picomole level (S/N = 3). This system successfully analyzed sequences of Leu(5)-enkephalin (25 pmol) and angiotensin I(100 pmol) using fluorometric detection at 524 nm with excitation at 387 nm.