Carboxyl-terminal protein sequencing method and kit
AssigneeThe Board of Trustees of the Leland Stanford Junior University
InventorsDean A. Le, Kazuhiko Tatemoto
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Abstract
Polypeptides are sequenced from the carboxyl terminus by treating the peptide with an iodoxybenzene, followed by removal of the excess reagent and reaction with 2-aminothiophenol at an elevated temperature. The resulting benzthiazolidine is then analyzed to determine the C-terminal amino acid which has been cleaved. A kit which contains the iodoxybenzene and 2-aminothiophenol reagents is separate containers is also disclosed.
Claims9
A method for carboxyl terminal sequencing of peptides, said method comprising:
- having protective groups on other available functionalities with an aryliodonium compound, whereby a geminaldiamino group is formed at the carboxyl terminus of the polypeptide;
- heating the polypeptide in the presence of a vicinal aminothiol to produce a thiazolidine having a side group derived from the carboxyl terminal amino acid of the polypeptide;
- and
- determining the composition of the thiazolidine, to determine the carboxyl terminal amino acid of said polypeptide.
A method for carboxyl terminal sequencing of peptides, said method comprising:
- bonding a sample polypeptide at its amino groups to an activated solid support;
- protecting other available functional groups other than carboxyl groups;
- activating the carboxyl group at the carboxyl terminus of the polypeptide and reacting the activated carboxyl group with glycinamide to provide a modified polypeptide having a terminal amide at its carboxyl terminus;
- reacting the modified polypeptide with an aryl iodonium compound, whereby a geminal-diamino group is formed at its carboxyl terminus;
- heating the modified polypeptide in the presence of o-aminothiophenol to produce a thiazolidine having a side group derived from the carboxyl terminal amino acid of the polypeptide;
- and
- determining the composition of the thiazolidine, to determine the carboxyl terminal amino acid of said polypeptide.
Dependencies are inferred from the claim text.