Peptide constructs and assay systems

Abstract

The present invention provides methods for constructing peptide construct sets and methods of use of these peptide construct sets in assay systems for peptide analysis, and in particular for use in high throughput peptide analysis. The methods allow for analysis of large sets of peptide constructs in a cost-effective manner, employing molecular biological techniques that are both robust and easily parallelized. Thus, the methods allow for the construction of peptide construct sets encompassing, e.g., the human proteome.

1 . A set of at least 5,000 distinct peptide constructs, each comprising a peptide portion and an identifying oligonucleotide portion, wherein the peptide portions are encoded by oligonucleotide sequences that are custom-designed so that at least 10% of the set of peptide constructs produced contain contiguous peptide sequences of at least 12 amino acids that have more than 80% amino acid identity to protein sequences encoded by up to 100 different species of organism. 
     
     
         2 . The set of peptide constructs from  claim 1 , where the species are eukaryotic species. 
     
     
         3 . The set of peptide constructs from  claim 2 , where the species are mammalian species. 
     
     
         4 . The set of peptide constructs from  claim 1 , where the species are bacterial species. 
     
     
         5 . The set of peptide constructs from  claim 1 , where the species are human pathogens. 
     
     
         6 . The set of peptide constructs from  claim 1 , where the peptide sequences are grouped into one or more sets of partially overlapping sequences. 
     
     
         7 . The set of peptides from  claim 1 , wherein the peptide portion is dissociated from the ribosome before being linked to the identifying nucleic acid. 
     
     
         8 . The set of peptide constructs from  claim 1 , wherein the set of peptide constructs comprises at least 100,000 distinct peptide constructs, each comprising a peptide portion of at least 10 amino acids and an identifying oligonucleotide portion, wherein the peptide portions are encoded within oligonucleotide sequences that are custom-designed such that at least 90% of the sequences contain no more than one in-frame codon that can function as a stop codon in the translation system used to make the peptide constructs. 
     
     
         21 . A method for analyzing a sample using the set of peptide constructs of  claim 1 , comprising the steps of:
 performing an assay on the set of peptide constructs, where some of the peptide constructs are acted upon by assay agents,   separating the peptide constructs that are acted upon by the assay agents from the peptide constructs that are not acted upon by the assay agents, and   determining the identities of substantially all of the peptide constructs of either the peptide constructs that were acted upon by the assay agents or the peptide constructs that were not acted upon by the assay agents by analyzing the nucleic acid portion of the peptide constructs.   
     
     
         22 . The method of  claim 21 , wherein the action of the assay agents results in some of the peptide constructs being modified permanently or non-permanently. 
     
     
         23 . The method of  claim 21 , wherein analyzing step is performed by digital sequencing. 
     
     
         24 . The method of  claim 21 , wherein analyzing step is performed by hybridization to a microarray. 
     
     
         25 . The method of  claim 21 , where at least 100,000 different peptide constructs are assayed. 
     
     
         26 . The method of  claim 21 , where at least 1,000,000 different peptide constructs are assayed.