Stereochemical control of DNA biosynthesis.

Abstract

Stereochemical control of DNA biosynthesis was studied using several DNA-synthesizing complexes containing, in each case, a single substitution of a 2'-deoxy-D-nucleotide residue by an enantiomeric L-nucleotide residue in a DNA chain (either in the primer or in the template) as well as 2'-deoxy-L-ribonucleoside 5'-triphosphates (L-dNTPs) as substrates. Three template-dependent DNA polymerases were tested, Escherichia coli DNA polymerase I Klenow fragment, Thermus aquaticus DNA polymerase and avian myeloblastosis virus reverse transcriptase, as well as template-independent calf-thymus terminal deoxynucleotidyl transferase. Very stringent control of stereoselectivity was demonstrated for template-dependent DNA polymerases, whereas terminal deoxynucleotidyl transferase was less selective. DNA polymerase I and reverse transcriptase catalyzed formation of dinucleoside 5',5'-tetraphosphates when L-dTTP was used as substrate. Comparison between models of template-primer complexes, modified or not by a single L-nucleotide residue, revealed striking differences in their geometry.