American Chemical Society
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Effects of Monomer and Template Concentration on the Kinetics of Nonenzymatic Template-Directed Oligoguanylate Synthesis

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journal contribution
posted on 1998-08-15, 00:00 authored by Anastassia Kanavarioti, Claude F. Bernasconi, Eldon E. Baird
To identify key parameters which influence the efficiency of nonenzymatic template-directed oligonucleotide reactions, a kinetic study of oligoguanylate synthesis on a polycytidylate (poly(C)) template has been performed. This oligomerization is satisfactorily described by three kinetic processes:  (i) dimerization to form the first primer (k2), (ii) elongation of a preformed primer (ki, i ≥ 3), and (iii) hydrolysis of the monomer to form deactivated material (kh), with kh < k2 < ki. This is the first reported study that includes rate determinations of ki as a function of the concentration of both poly(C) template and the activated monomer, guanosine 5‘-monophosphate-2-methylimidazolide (2-MeImpG), in the range 2 mM ≤ [poly(C)] ≤ 50 mM and 5 mM ≤ [2-MeImpG] ≤ 50 mM. ki values determined under conditions where the template is fully saturated with monomer are practically independent of both monomer and polymer concentration and thus strongly support a template-directed elongation model. Values of ki determined with a partially occupied template support a mechanism wherein the reaction of the oligonucleotide primer with a template-bound monomer is assisted by the presence of two additional downstream template-bound 2-MeImpG molecules. Comparison between the kinetic parameters obtained here and the ones determined in the montmorillonite-catalyzed oligoadenylate polymerization allows the proposition that the ratio of the rate constants ki/kh determines efficiency and the ratio ki/k2 determines the degree or extent of a polymerization. These conclusions provide new design principles for the optimization of nonenzymatic polymerizations.