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2008 Annual Science Report

University of Colorado, Boulder Reporting  |  JUL 2007 – JUN 2008

A Novel Route to New, Simpler, Self-Aminoacylating Ribozymes

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A Novel Route to New, Simpler, Self-aminoacylating Ribozymes

Selection-amplification finds new RNA enzymes (ribozymes) among randomized RNAs with flanking fixed sequences (primer complements). Precise removal of 3’-primer before aminoacylation selected aminoacylation from PheAMP in 3 cycles, yielding active transfer (kcat = 12-20 min-1), strikingly using only three conserved nucleotides, and acting independently of divalent ions. This unusually simple RNA active site encouraged study of the reaction via molecular mechanics-based free energy minimization. On this basis, we can suggest a chemical path for RNA-catalyzed transacylation. The site model also predicted new features — L-stereoselectivity, 2’-regioselectivity, independence of amino acid side chain and phosphorylated activating group, which were verified. The selection’s outcome also indicates that RNA aminoacylation from adenylate is simpler than aminoacylation from CoA thioester, potentially accounting for the exclusive translational use of adenylates in biological evolution. The simplicity of this active site suggests a general route to small ribozymes, which may have been evolved via evolutionary processes beginning with small-nonspecific active centers.

- N. V. Chumachenko, Y. Novikov, M. Yarus, MCD Biology, University of Colorado Boulder