Templating efficiency of naked DNA.
Template-directed synthesis of complementary strands is pivotal for life. Nature employs polymerases for this reaction, leaving the ability of DNA itself to direct the incorporation of individual nucleotides at the end of a growing primer difficult to assess. Using 64 sequences, we now find that any of the four nucleobases, in combination with any neighboring residue, support enzyme-free primer extension when primer and mononucleotide are sufficiently reactive, with >or=93% primer extension for all sequences. Between the 64 possible base triplets, the rate of extension for the poorest template, CAG, with A as templating base, and the most efficient template, TCT, with C as templating base, differs by less than two orders of magnitude. Further, primer extension with a balanced mixture of monomers shows >or=72% of the correct extension product in all cases, and >or=90% incorporation of the correct base for 46 out of 64 triplets in the presence of a downstream-binding strand. A mechanism is proposed with a binding equilibrium for the monomer, deprotonation of the primer, and two chemical steps, the first of which is most strongly modulated by the sequence. Overall, rates show a surprisingly smooth reactivity landscape, with similar incorporation on strongly and weakly templating sequences. These results help to clarify the substrate contribution to copying, as found in polymerase-catalyzed replication, and show an important feature of DNA as genetic material.
Kervio
Eric
E
Institut für Organische Chemie, Universität Stuttgart, 70569 Stuttgart, Germany.
Hochgesand
Annette
A
Steiner
Ulrich E
UE
Richert
Clemens
C
D001483:Base Sequence; D004247:DNA / Q000096:biosynthesis / Q000737:chemistry / Q000235:genetics*; D017931:DNA Primers / Q000235:genetics*; D004261:DNA Replication; D008956:Models, Chemical; D008957:Models, Genetic; D015394:Molecular Structure; D013698:Templates, Genetic
D017931:DNA Primers; D004247:DNA