ORGN 653 |
| An ongoing challenge for deoxyribozyme-mediated RNA ligations has been synthesis of native 3'-5' phosphodiester linkages. Previously, reactions of 5'-triphosphate RNA with a second RNA substrate have provided mostly 2',5'-branched RNA. Here we report two new strategies designed to overcome this problem. Both strategies utilize in vitro selection to identify Mg2+-dependent deoxyribozymes that synthesize 3'-5'-linked linear RNA from a 2',3'-diol and a 5'-triphosphate. The first strategy controls regioselectivity by placing the incipient linkage within a DNA:RNA duplex region, such that 3'-5' is favored over 2'-5' both thermodynamically and kinetically. The second strategy makes use of a known deoxyribozyme that reacts only with 3'-5' RNA linkages. Incorporating this separate deoxyribozyme as part of an additional step in the selection cycle allows isolation of new deoxyribozymes that prepare only native linkages. We present data on the characterization of RNA ligation activity and the generality of the new DNA enzymes from both ligation strategies.
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Proteins, Peptides, and Nucleotides
8:00 AM-11:00 AM, Wednesday, 16 March 2005 Convention Center -- Room 9, Oral
Division of Organic Chemistry |
