New methods and kits for sequencing polypeptides
AssigneeThe Procter & Gamble Company
InventorsThomas Keough, Robert Youngquist
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Abstract
The present disclosure provides methods and kits which are useful for sequencing polypeptides. The methods involve derivatization of the N-terminus of the polypeptide or peptides thereof. The methods also involve derivatization of the epsilon amino group of the side-chain of the lysine containing polypeptide or peptides thereof. Mass spectral analysis of one or more of the resulting derivatized analytes provides spectra which are readily interpreted through the use of techniques well-known to the ordinarily skilled artisan. The present disclosure also describes kits which enhance convenient performance of the methods.
Claims38
A method of determining the amino acid sequence of a polypeptide wherein one or more peptides of the polypeptide contain lysine, said method comprising:
- (a) converting the epsilon amino group of lysine on the lysine containing peptides to a guanine or another very basic group, a fixed cationic-charged group or an isotopically labeled group;
- (b) derivatizing the N-terminus of the polypeptide or the N-termini of one or more peptides of the polypeptide with one or more acidic moieties having pKas of less than about 2 when coupled with the polypeptide or peptides, to provide one or more derivatized analytes;
- (c) analyzing one or more derivatized analytes using a mass spectrometric technique to provide a fragmentation pattern;
- and
- (d) interpreting the fragmentation pattern.
A kit for use in determining the amino acid sequence of a polypeptide wherein one or more peptides of the polypeptide contain lysine, said kit comprising:
- (a) one or more chemical reagents for converting the epsilon amino group of lysine on the lysine containing peptides to a guanine or other basic group or fixed cationic-charge group or an isotopically labeled group;
- (b) means for converting the epsilon amino group of lysine on the lysine containing peptides to a guanine or other basic group or fixed cationic-charge groups or an isotopically labeled group with said chemical reagents;
- (c) one or more acidic moiety reagents providing one or more acidic moieties having pKas of less than about 2 when coupled with the polypeptide or one or more peptides of the polypeptide;
- and
- (d) means for derivatizing the N-terminus of the polypeptide or the N-termini of one or more peptides of the polypeptide with one or more acidic moiety reagents.
Dependencies are inferred from the claim text.