Chemical carboxyl-terminal sequence analysis of peptides and proteins using tribenzylsilyl isothiocyanate.
A new derivatization reagent, tribenzylsilyl isothiocyanate (TBS-ITC), is applied to C-terminal peptide and protein sequencing. It has been successfully used to sequence six C-terminal residues of house apomyoglobin and a synthetic peptide at low nanomole levels. The chemistry involves activation with acetic anhydride, derivatization with TBS-ITC, and cleavage of derivatized C-terminal amino acid thiohydantoin with sodium hydroxide. The tribenzylsilyl is a bulky, electric donor group and is a good leaving group. It facilitates the nucleophilic attack of the NCS(-1) in the coupling reaction. The efficiency for C-terminal sequencing by TBS-ITC is about the same as that of acetyl isothiocyanate (AITC), which is a derivatizing reagent for C-terminal sequencing developed by our laboratory. TBS-ITC is much more stable than AITC and trimethylsilyl isothiocyanate (TMS-ITC). TBS-ITC is a solid with relatively long shelf life, whereas AITC and TMS-ITC are liquid and not stable at room temperature.
Liu
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Y
College of Life Sciences, Hunan Normal University, Changsha, People's Republic of China.
Liang
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S
D000595:Amino Acid Sequence; D000596:Amino Acids / Q000032:analysis / Q000737:chemistry; D000818:Animals; D001059:Apoproteins / Q000737:chemistry; D002851:Chromatography, High Pressure Liquid; D006736:Horses; D017879:Isothiocyanates / Q000737:chemistry*; D013058:Mass Spectrometry; D009211:Myoglobin / Q000737:chemistry; D010455:Peptides / Q000737:chemistry*; D011506:Proteins / Q000737:chemistry*; D020539:Sequence Analysis, Protein / Q000379:methods*; D013055:Spectrophotometry, Infrared; D013867:Thiohydantoins / Q000032:analysis / Q000737:chemistry
D000596:Amino Acids; D001059:Apoproteins; D017879:Isothiocyanates; D009211:Myoglobin; D010455:Peptides; D011506:Proteins; D013867:Thiohydantoins; C010857:apomyoglobin; C461061:tribenzylsilyl isothiocyanate