Method of preparing a protein array based on biochemical protein-protein interaction

Abstract

A method of preparing a protein array based on biochemical protein-protein interaction is provided. An array of a first protein which comprises a PDZ domain is deposited on a substrate. A second protein, which comprises an amino acid sequence (S/T)-X-(V/I/L)-COOH (each hyphen represents a peptide bond, each parenthesis encloses amino acids which are alternatives to one other, each slash within such parentheses separates the alternative amino acids, and the X represents any amino acid which is selected from the group comprising the twenty naturally occurring amino acids), is applied to the first protein array. The amino acid sequence (S/T)-X-(V/I/L)-COOH of the second protein is bound to the PDZ domain of the first protein.

1 . A method of preparing a protein array based on biochemical protein-protein interaction, comprising the steps of: 
 (a) depositing on a substrate an array of a first protein, the first protein comprising a PDZ domain; and    (b) applying a second protein, which comprises an amino acid sequence (S/T)-X-(V/I/L)-COOH, to the first protein array, the amino acid sequence (S/T)-X-(V/I/L)-COOH of the second protein binding to the PDZ domain of the first protein,    wherein each hyphen represents a peptide bond, each parenthesis encloses amino acids which are alternative to one other, each slash within such parentheses separates the alternative amino acids, and the X represents any amino acid which is selected from the group consisting of alanine, cysteine, aspartic acid, glutamic acid, phenylalanine, glycine, histidine, isoleucine, lysine, leucine, methionine, asparagine, proline, glutamine, arginine, serine, threonine, valine, tryptophan and tyrosine.    
     
     
         2 . The method of  claim 1 , wherein the amino acid sequence (S/T)-x-(V/I/L) is fused to the C-terminal of the second protein.  
     
     
         3 . The method of  claim 1 , wherein the protein array is maintained under physiological condition, and is used to screen one or more drug targets.  
     
     
         4 . The method of  claim 1 , wherein the first protein deposited in step (a) is in a soluble buffer.  
     
     
         5 . The method of  claim 1 , wherein the first protein deposited in step (a) is immobilized in a gel.  
     
     
         6 . The method of  claim 1 , wherein the substrate includes a plurality of microwells contained therein, and the first protein is deposited in step (a) into the microwells.  
     
     
         7 . The method of  claim 1 , wherein the substrate includes a glass plate, and the first protein array is printed onto the glass plate in step (a).  
     
     
         8 . The method of  claim 1 , wherein the substrate includes a glass plate and a plurality of gel pads on the glass plate, and the first protein is deposited in step (a) onto the gel pads.  
     
     
         9 . The method of  claim 1 , wherein the first protein is deposited on the substrate by a robot.  
     
     
         10 . The method of  claim 1 , wherein at least one array element of the protein array includes an oligonucleotide.  
     
     
         11 . The method of  claim 1 , wherein at least one array element of the protein array includes messenger RNA.  
     
     
         12 . The method of  claim 1 , wherein at least one array element of the protein array includes DNA.  
     
     
         13 . The method of  claim 1 , wherein at least one array element includes a sugar.  
     
     
         14 - 15  (canceled).  
     
     
         16 . A method of preparing a protein polypeptide array, comprising the steps of: 
 (a) depositing on a substrate an array of a first polypeptide, the first polypeptide comprising a PDZ domain; and    (b) applying a second polypeptide which comprises an amino acid sequence (s/T)-X-(V/I/L)-COOH to the first polypeptide array, the amino acid sequence (S/T)-X-(V/I/L)—COOH of the second polypeptide binding to the PDZ domain of the first polypeptide,    wherein each hyphen represents a peptide bond, each parenthesis encloses amino acids which are alternatives to one other, each slash within such parentheses separates the alternative amino acids, and the X represents any amino acid which is selected from the group consisting of alanine, cysteine, aspartic acid, glutamic acid, phenylalanine, glycine, histidine, isoleucine, lysine, leucine, methionine, asparagine, praline, glutamine, arginine, serine, threonine, valine, tryptophan and tyrosine.    
     
     
         17 . The method of  claim 16 , wherein at least one array element includes an oligonucleotide in addition to the first polypeptide.  
     
     
         18 . The method of  claim 16 , wherein at least one array element includes messenger RNA in addition to the first polypeptide.  
     
     
         19 . The method of  claim 16 , wherein at least one array element includes DNA in addition to the first polypeptide.  
     
     
         20 . The method of  claim 16 , wherein at least one array element includes a sugar in addition to the first polypeptide.