A complete quantitative N-terminal analysis method.
A complete quantitative N-terminal analysis (QNA) technique based on the application of dimethylaminoazobenzene isothiocyanate is described. The method allows recovery of all free N-terminal amino acids, including Asn, Gln, Trp, Ser, and Thr in quantitative yield. N-Termini of polypeptides as little as 5 pmol can be reliably and reproducibly determined by this method. This QNA method is useful in many aspects of protein structure analysis. (a) QNA is useful in assessing the purity, identity, and quantity of a polypeptide preparation. It has also been applied in our lab as a routine guarding step to prevent impure or ill-characterized samples from occupying the space of the gas-phase sequenator. (b) QNA of a trypsinized protein generates a miniaturized amino acid composition which is useful both in characterizing the identity of a protein and in comparing the homology of structurally related proteins. (c) QNA can be used to follow the pathway and preferential cleavage sites of limited proteolysis. (d) QNA is useful in characterizing selectively modified Lys and Arg residues. The details of this QNA method and the results of its applications are presented here.
D000596:Amino Acids / Q000032:analysis; D002851:Chromatography, High Pressure Liquid; D006801:Humans; D017879:Isothiocyanates; D008832:Microchemistry; D010449:Peptide Mapping; D010455:Peptides / Q000032:analysis*; D011487:Protein Conformation; D013861:Thiocyanates; D004124:p-Dimethylaminoazobenzene / Q000031:analogs & derivatives*
D000596:Amino Acids; D017879:Isothiocyanates; D010455:Peptides; D013861:Thiocyanates; C012926:4-(N,N-dimethylaminoazobenzene)-4'-isothiocyanate; D004124:p-Dimethylaminoazobenzene