Methods and compositions for protein and peptide sequencing

A method of sequencing a peptide, comprising:

• (a) incubating the peptide with a library of DNA aptamers exhibiting binding specificity toward at least one N-terminal amino acid under conditions where one or more aptamers bind specifically to at least one N-terminal amino acid of the peptide, wherein each aptamer in the library comprises a peptide binding ssDNA region and a unique barcode sequence indicative of the first binding round and the associated peptide binding ssDNA region;
• (b) ligating the DNA aptamer bound to the N-terminal of the peptide onto a proximal DNA barcode construct;
• (c) removing the peptide binding sequence from the DNA aptamer, thereby leaving the barcode of the DNA aptamer covalently attached to the DNA barcode construct:
• (d) removing the N-terminal amino acid from the peptide to produce an N-terminal amino acid-shortened peptide;
• (e) incubating the N-terminal amino acid-shortened peptide with the library of aptamers exhibiting binding specificity toward at least one N-terminal amino acid under conditions where one or more aptamers bind specifically to at least one N-terminal amino acid of the N-terminal amino acid-shortened peptide, wherein each aptamer in the library comprises a peptide binding ssDNA region and a unique barcode sequence indicative of the second binding round and the associated peptide binding ssDNA region;
• (f) ligating the DNA aptamer bound to the N-terminal of the peptide onto the proximal DNA barcode construct;
• (g) removing the peptide binding sequence from the DNA aptamer, thereby leaving the barcode of the DNA aptamer covalently attached to the DNA barcode construct;
• (h) removing the N-terminal amino acid from the N-terminal amino acid-shortened peptide;
• (i) repeating steps (a)-(d) a plurality of times to construct a chain of positional barcodes that correspond to sequential N-terminal amino acids in the peptide;
• and
• (j) sequencing the chain of positional barcodes to thereby obtain the sequence of the peptide.