An automated fluorescence protein sequencer using 7-methylthio-4-(2,1,3-benzoxadiazolyl) isothiocyanate (MTBD-NCS) as an Edman reagent.
An automated fluorescence protein sequencer using 7-methylthio-5-(2,1,3-benzoxadiazolyl) isithiocyanate (MTBD-NCS), a fluorescent Edman reagent, is developed by the modification of a commercial protein sequencer. The generated MTBD-thiohydantoin amino acids fluoresced strongly, whereas the by-products such as MTBD-thiocarbamoyl amino acids and MTBD-carbamoly amino acids did not fluoresce. A few interfering peaks were observed in the chromatogram and amino acid sequence was easily determined. The coupling and cyclization/cleavage reaction conditions and extraction conditions of generated MTBD-thiazolinone amino acids were optimized using an autonalyzer. Finally, the sequence of a synthetic peptide (25 pmol), leucine-enkephalin-Thr-amide, was determined and up to six residues were successively analyzed.
Okiyama
Natsuko
N
Graduate School of Pharmaceutical Sciences, The University of Tokyo, 7-3-1 Hongo, Bunkyo-ku, Tokyo 113-0033, Japan.
Santa
Tomofumi
T
Toriba
Akira
A
Nakagomi
Kazuya
K
Imai
Kazuhiro
K
Hiranuma
Hidenori
H
Tanaka
Hiroshi
H
D005456:Fluorescent Dyes / Q000737:chemistry*; D017879:Isothiocyanates / Q000737:chemistry*; D010069:Oxadiazoles / Q000737:chemistry*; D011506:Proteins / Q000737:chemistry*; D020539:Sequence Analysis, Protein; D013050:Spectrometry, Fluorescence
C406762:7-methylthio-4-(2,1,3-benzoxadiazolyl)isothiocyanate; D005456:Fluorescent Dyes; D017879:Isothiocyanates; D010069:Oxadiazoles; D011506:Proteins