Stereoselective Synthesis using Immobilized Diels–Alderase Ribozymes

Abstract

Development of artificial ribozymes by in vitro selection has so far, mostly been addressed from the viewpoint of fundamental research. However, such ribozymes also have high potential as selective catalysts in practical syntheses. Immobilization of an active and selective ribozyme is an important step towards this end. A 49-nucleotide RNA molecule that was previously found to stereoselectively catalyze Diels–Alder reactions between various anthracene dienes and maleimide dienophiles was quantitatively immobilized on an agarose matrix by periodate oxidation of the 3′-terminal ribose and coupling to a hydrazide moiety. Typical loadings were 45 pmol μL⁻¹ gel. The specific activity was comparable to that of soluble ribozyme, and high enantioselectivities were obtained in catalyzed cycloadditions. The catalytic matrix was found to be stable and could be regenerated about 40 times with only minimal reduction of catalytic activity. Like the soluble ribozyme, the immobilized catalyst stereoselectively converts various diene and dienophile substrates. By using either natural D-RNA or enantiomeric L-RNA, both product enantiomers were made synthetically accessible with similar selectivities.