Peptide microarray and method of use
View Patent ↗A peptide microarray comprising a plurality of predicted unique binding peptides being selected by computational prediction of interaction with a protein of interest or a domain thereof. The selected unique binding peptides are pre-synthesized and then printed and/or immobilized onto a solid support surface via N-terminus with a linker. Methods of using the invention peptide microarray for quantitative determination of protein-peptide interaction, epitope mapping, and drug screening are also provided.
1. A method of identifying one or more binding peptides interacting with a protein of interest or a domain thereof, wherein at least one binding peptide or the protein of interest is methylated or acetylated, said method comprising:
a) providing computational predictions for predicted unique binding peptides to the protein of interest or a domain thereof;
b) selecting a plurality of predicted unique binding peptides based on the computational predictions;
c) generating a peptide microarray comprising the plurality of the predicted unique binding peptides being pre-synthesized and capped with alanine at each terminus and immobilized on a solid support surface by a linker comprising aminohexanoic acid (Ahx) or polyethylene glycol (PEG), said alanine cap at each terminus stabilizing the immobilization on the solid support surface;
d) exposing the microarray to the protein of interest or domain thereof; and
e) detecting interactions of predicted binding peptides with the protein of interest or domain thereof on the microarray, thereby identifying one or more binding peptides interacting with a protein of interest or a domain thereof.
2. The method of claim 1 , wherein said computational predictions comprises a combination of at least three analytical approaches selected from the group consisting of structural information, energetic pattern of said peptide-protein interactions, molecular dynamics simulation, conservation during evolution, mutation, subcellular location, functional filtering processes, proteome scan, and mass spectra evidence.
3. The method of claim 1 , wherein between 100-1000 predicted unique binding peptides are immobilized on the solid support surface.
4. The method of claim 1 , wherein the linker comprises polyethylene glycol (PEG).
5. The method of claim 1 , wherein at least one predicted unique binding peptide is methylated.
6. The method of claim 1 , wherein said protein of interest or a domain thereof is methylated.
7. The method of claim 1 , wherein said detecting comprises detection of a fluorescent signal that correlates with a binding affinity and provides a quantitative measurement of interaction between said binding peptides and said protein of interest or protein domain thereof.
8. The method of claim 1 , wherein said binding peptide-protein interaction is a transient interaction.
9. The method of claim 1 , wherein said linker comprises aminohexanoic acid (Ahx).
10. The method of claim 1 , wherein at least one predicted unique binding peptide is acetylated.
11. The method of claim 1 , wherein said protein of interest or a domain thereof is acetylated.