Chromatographic resolution of two DNA polymerase activities from bloodstream forms of [formula omitted]: Differential responses to exogenous polyamine addition

Abstract

A single peak of DNA polymerase activity from extracts of T. brucei , obtained by DEAE-cellulose and phosphocellulose ion-exchange chromatography, was resolved into two peaks differing in KCl concentration necessary to elute them from a DNA-agarose column. Peak I (eluting at 0.2 M KCl) and Peak II (eluting at 0.4 M KCl), differed in response to increasing KCl concentrations, although both functioned optimally with Mg 2+ as divalent cation when DNA synthesis was directed either by activated DNA or poly (dC)·(dG) 12–18. Due to the potential significance of polyamines in the metabolism of T. brucei , the effect of exogenous polyamine on rates of DNA synthesis by the peak I and II enzymes was compared with that of murine DNA polymerase alpha. Only the peak I enzyme was significantly stimulated (up to 4-fold) by the biologically active polyamines spermine and spermidine at physiological concentrations. The response of the peak I enzyme resembled that of the alpha polymerase. This result suggests a possible functional difference between peak I and II enzymes, as well as a potential target site for trypanocidal drug development.