Isotope distribution encoded tags for protein identification

A method for identifying a protein in a mixture of proteins that is at least in part encoded by a known expressed sequence tag which comprises the steps of:

• (a) labeling a protein in the mixture with a first isotope-distribution tag reagent which comprises an element having an isotope distribution not normally found in the proteins of the mixture and which selectively reacts and bonds to a functional group in a protein to give a labeled protein;
• (b) digesting the labeled protein using a digestion method that is selective for a cut site in the protein to generate digested peptides including peptides that are selectively labeled with the first isotope-distribution tag reagent derived from proteins in the labeled protein mixture;
• (c) measuring the masses of the digested peptides selectively labeled with the first isotope-distribution tag reagent employing mass spectrometry to give the masses of the digested labeled peptides;
• and
• (d) comparing the mass of one and only one single digested labeled peptide determined in step (c) with one or more databases comprising the masses of peptides encoded by known expressed sequence tags to identify peptides in the database that correspond to the single digested peptide and thereby determine which protein(s) in the mixture are encoded at least in part by a given expressed sequence tag.

A method for identifying a peptide or protein in a mixture comprising the steps:

• (a) labeling one or more peptides or proteins in a mixture with an isotope-distribution tag reagent that selectively labels one or more amino acids in one or more peptides or proteins in the mixture to give one or more labeled peptides or proteins;
• (b) determining the masses of the labeled peptides or proteins by mass-spectrometry to give the masses of the peptides or proteins;
• and
• (c) identifying a peptide or protein in a database using the mass of one and only one single labeled peptide or protein and a sequence constraint derived from the specificity of the first isotope-distribution tag reagent;
• wherein the mass of the single peptide or protein and the sequence constraint identifies a peptide or protein in the database.

A method for identifying a peptide or protein in a mixture comprising the steps:

• (a) labeling one or more peptides or proteins in a mixture with an isotope-distribution tag reagent that selectively labels one or-more uncommon amino acids in one or more peptides or proteins in the mixture to give one or more labeled peptides or proteins;
• (b) digesting the mixture using a digestion method that is selected to generate one or more digested labeled peptides or proteins;
• (c) determining the masses of said digested labeled peptides or proteins by mass-spectrometry to give the masses of the digested labeled peptides or proteins;
• and
• (d) identifying a protein or peptide in a database using the mass of one and only one single digested labeled peptide or protein and a sequence constraint derived from the specificity of the first isotope-distribution tag reagent;
• wherein the mass of the single digested labeled peptide or protein and the sequence constraint identifies a peptide or protein in the database.

A method for identifying a peptide or protein in a mixture comprising the steps:

• (a) labeling one or more peptides or proteins in a mixture with an isotope-distribution tag reagent that selectively labels one or more amino acids in one or more peptides or proteins in the mixture to give one or more labeled peptides or proteins;
• (b) fragmenting the peptides or proteins in the mixture in a mass spectrometer to generate one or more fragmented peptides or proteins selectively labeled with an isotope-distribution tag reagent;
• (c) determining the masses of said fragmented peptides or proteins by mass-spectrometry to give the masses of one or more fragmented peptides or proteins selectively labeled with an isotope-distribution tag reagent;
• and
• (d) identifying a protein or peptide in a database using the mass of one and only one single fragmented peptide or protein selectively labeled with an isotope-distribution tag reagent and a sequence constraint derived from the specificity of the first isotope-distribution tag reagent and without determining the sequence of any ion derived from the one or more digested labeled peptides or proteins;
• wherein the mass of the single fragmented peptide or protein and the sequence constraint identifies a peptide or protein in the database.