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Identification of Small Molecule Binding Sites within Proteins Using Phage Display Technology

[ Vol. 4 , Issue. 7 ]

Author(s):

D. J. Rodi, G. E. Agoston, R. Manon, R. Lapcevich, S. J. Green and L. Makowski   Pages 553 - 572 ( 20 )

Abstract:


Affinity selection of peptides displayed on phage particles was used as the basis for mapping molecular contacts between small molecule ligands and their protein targets. Analysis of the crystal structures of complexes between proteins and small molecule ligands revealed that virtually all ligands of molecular weight 300 Da or greater have a continuous binding epitope of 5 residues or more. This observation led to the development of a technique for binding site identification which involves statistical analysis of an affinity-selected set of peptides obtained by screening of libraries of random, phage-displayed peptides against small molecules attached to solid surfaces. A random sample of the selected peptides is sequenced and used as input for a similarity scanning program which calculates cumulative similarity scores along the length of the putative receptor. Regions of the protein sequence exhibiting the highest similarity with the selected peptides proved to have a high probability of being involved in ligand binding. This technique has been employed successfully to map the contact residues in multiple known targets of the anticancer drugs paclitaxel (Taxol™), docetaxel (Taxotere™) and 2-methoxyestradiol and the glycosaminoglycan hyaluronan, and to identify a novel paclitaxel receptor [1]. These data corroborate the observation that the binding properties of peptides displayed on the surface of phage particles can mimic the binding properties of peptides in naturally occurring proteins. It follows directly that structural context is relatively unimportant for determining the binding properties of these disordered peptides. This technique represents a novel, rapid, high resolution method for identifying potential ligand binding sites in the absence of three-dimensional information and has the potential to greatly enhance the speed of development of novel small molecule pharmaceuticals.

, Identification, of, Small, Molecule, Binding, Sites, within, Proteins, Using, Phage, Display, Technology

Affiliation:

Biosciences Division, Argonne National Laboratory, 9700 South Cass Avenue, Argonne, IL 60439, USA.



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