Content area
Full Text
Key Words DNA polymerase, replication fidelity, Watson-Crick hydrogen bonding
* Abstract Understanding the mechanisms by which genetic information is replicated is important both to basic knowledge of biological organisms and to many useful applications in biomedical research and biotechnology. One of the main functions of a DNA polymerase enzyme is to help DNA recognize itself with high specificity when a strand is being copied. Recent studies have shed new light on the question of what physical forces cause a polymerase enzyme to insert a nucleotide into a strand of DNA and to choose the correct nucleotide over the incorrect ones. This is discussed in the light of three main forces that govern DNA recognition: base stacking, Watson-Crick hydrogen bonding, and steric interactions. These factors are studied with natural and structurally altered DNA nucleosides.
INTRODUCTION
The sequence-specific polymerization of DNA is central to the existence of living organisms on Earth. The chemistry of this reaction is simple: the 3' hydroxyl group on the end of an existing primer strand forms a phosphodiester with a 5' phosphate on a nucleotide, displacing a pyrophosphate leaving group (15, 30-32). The thermodynamic driving force is the cleavage of the weaker phosphatephosphate bond coupled with formation of a stronger phosphodiester bond and the entropy of freeing the pyrophosphate. In principle, this reaction can occur without an enzyme, with only DNA and free nucleotides; however, it is exceedingly slow, whereas DNA polymerase enzymes can perform this at the rate of several hundred times per second. The enzyme speeds this reaction by lowering the activation barrier for the reaction by several kilocalories per mole.
The function of a DNA polymerase is to form base pairs from an existing single template strand and free nucleotides in solution, and to do so with high specificity for forming correct pairs rather than incorrect ones (15, 30-32). Thus, a polymerase helps DNA recognize itself with high specificity. To best understand how polymerases do this, it is useful first to consider how DNA alone recognizes itself, and then to understand how the enzyme influences this recognition.
We focus on what is currently understood about the forces that govern DNADNA recognition in the absence and presence of polymerase enzymes. First I briefly discuss each of the...