RNA aptamers directed to discrete functional sites on a single protein structural domain

摘要: Cellular regulatory networks are organized such that many proteins have few interactions, whereas a many. These densely connected protein “hubs” critical for the system-wide behavior of cells, and capability selectively perturbing subset interactions at these hubs is invaluable in deciphering manipulating mechanisms. SELEX-generated RNA aptamers proving to be highly effective reagents inhibiting targeted proteins, but conventional methods generate one or several aptamer clones usually bind single target site most preferred by nucleic acid ligand. We advance generalized scheme isolating multiple sites on molecule reducing ability select its cognate aptamer. demonstrate use this generating directed discrete functional surfaces yeast TATA-binding (TBP). Previously we selected “class 1” interfere with TBP's binding TATA-DNA. By masking TBP TATA-DNA an unamplifiable class 1 aptamer, isolated new class, 2,” can TBP·DNA complex competition another general transcription factor, TFIIA. Moreover, show both inhibit polymerase II-dependent transcription, analysis template-bound factors shows they do so mechanistically distinct unexpected ways attributed either DNA TFIIA recognition sites. results should spur innovative approaches modulating other proteins.