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Unformatted text preview: Functionally Orthogonal Ligand- Receptor Pairs for the Selective Regulation of Gene Expression Generated by Manipulation of Charged Residues at the Ligand- Receptor Interface of ER r and ER Youheng Shi and John T. Koh* Contribution from the Department of Chemistry and Biochemistry, Uni V ersity of Delaware, Newark, Delaware 19716 Received August 20, 2001. Revised Manuscript Received January 3, 2002 Abstract: The reengineering of protein-small molecule interfaces represents a powerful tool of chemical biology. For many applications it is necessary to engineer receptors so that they do not interact with their endogenous ligands but are highly responsive to designed ligand analogues, which in turn do not interact with endogenous proteins. The chemical design strategy used to reengineer protein- small molecule interfaces is particularly challenging for interfaces involving relatively plastic receptor binding sites and therefore presents a unique challenge in molecular design. In this study we explore the scope and limita- tions of a new strategy for manipulating polar/charged residues across the ligand receptor interface of estradiol (E2) and the estrogen receptor (ER). Carboxylate-functionalized E2 analogues can activate ER R (Glu353 f Ala) and ER (Glu305 f Ala) with very large selectivites, demonstrating that this design strategy is extendable to other members of the steroid hormone receptor family. Neutral E2 analogues were found to complement ER R (E353A) with similar potencies but with generally lower selectivities. This suggests that the high selectivity observed with ligand- receptor pairs generated by exchanging charged residues across ligand- receptor interfaces is only due in part to their complementary shapes and that appropriate introduction of charged functionality on the ligand can provide substantial enhancement of selectivity by decreasing the engineered ligands affinity for the endogenous receptor. Attempts to modify the cationic residues by complementing Arg394 f Ala or Arg394 f Glu were not successful. Introduction The ability to create ligands that can selectively bind and activate engineered proteins represents one of the most important tools of chemical biology. 1 Of particular interest has been the creation of enzyme- substrate or ligand- receptor pairs capable of selectively controlling signal transduction events or gene transcription. There are numerous examples of engineered proteins that uniquely respond to substrate or ligand analogues; however, there are relatively few examples of engineered ligand receptor pairs that can truly function independently of the endogenous systems, having ligand analogues that do not react with the endogenous proteins and engineered receptors that do not interact with endogenous ligands. Thus, the exploration of new design strategies to engineer potent and functionally orthogonal ligand- receptor pairs represents an important chal- lenge in chemical design that would have a broad range of potential applications and may form the basis for developing...
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This note was uploaded on 12/04/2011 for the course CHEM 590A taught by Professor Staff during the Summer '10 term at University of Illinois, Urbana Champaign.
- Summer '10