Sequencing of peptides and proteins from the carboxy terminus.
A new chemical method for carboxy-terminal (C-terminal) protein sequencing has been developed. This approach has been successfully used to sequence 5 residues of standard proteins and 5 to 10 residues of synthetic peptides at low nanomole levels. The sequencing procedure consists of converting the C-terminal amino acid into a thiohydantoin (TH) derivative, followed by transformation of the TH into a good leaving group by alkylation. Next, the alkylated TH is cleaved mildly and efficiently with (N = C V S)- anion, which simultaneously forms a TH on the newly truncated protein or peptide. Thus, after the initial TH derivatization, there is no return to a free carboxyl group at the C-terminus. An additional benefit of this method is that the alkylating moiety can be chosen with a variety of properties allowing for variation in the detection method. This chemistry has been adapted to automated protein sequencers with a cycle time of about 1 h.
Boyd
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VL
Applied Biosystems, Inc., Foster City, California 94404.
Bozzini
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M
Zon
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G
Noble
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Mattaliano
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RJ
D000595:Amino Acid Sequence; D002851:Chromatography, High Pressure Liquid / Q000379:methods; D007202:Indicators and Reagents; D009682:Magnetic Resonance Spectroscopy / Q000379:methods; D010455:Peptides / Q000737:chemistry*; D011506:Proteins / Q000737:chemistry*
D007202:Indicators and Reagents; D010455:Peptides; D011506:Proteins