DNA Polymerase I enzyme biochemistry reflects life cycles of environmental phages

• Sydney Iredell, Pre-Veterinary Medicine and Animal Biosciences, University of Delaware

• Shawn Polson, Computer & Information Sciences, University of Delaware

Viruses are the most abundant and diverse biological entities on the planet. While viruses lack a universal marker gene, their replication genes allow researchers to explore viral diversity. Family A DNA Polymerase I (polA) is present in 25% of dsDNA phage with the sole function of genome replication. Prior studies have shown the 762 residue (Escherichia coli numbering) within the polA is responsible for dNTP incorporation, and substitutions at this site have been shown to alter the fidelity, speed and accuracy of the enzyme. Further research into bacteriophage polA suggests that this position will indicate phage life cycle. Amino acids Phenylalanine, Tyrosine and Leucine are the most common residues found at position 762 in phages. Three novel DNA Polymerase I enzymes (F_R2, Y_R3 and L1_S67) isolated from reference phages and metagenomic sequences were chosen to investigate the relationship between enzyme biochemistry and phage life cycle. Purified protein stocks were assessed for a range of in vitro biochemical parameters including purity, primer extension activity, and specific activity (speed). Results suggest that the range of temperatures that the enzymes are active generally reflects environmental temperature conditions, and the speed of the polymerase stocks closely reflect the known life history strategies (or lifecycles) of the phages from which they were isolated. Future work will include a measure of mutation rate for each polymerase.