KLENOW FRAGMENT AND DNA-POLYMERASE ALPHA-PRIMASE FROM SERVA CALF THYMUS IN WATER-IN-OIL MICROEMULSIONS

The activity of DNA polymerase alpha-primase complex from calf thymus and Klenow fragment of E. coli DNA polymerase 1 has been studied in re verse microemulsions formed by sodium bis(2-ethylhexyl) sulfosuccinate (AOT), sodium dodecylsulfate (SDS), cetyl trimethyl ammonium bromide (CTAB), polyoxyethylene 20 cetyl ether (Brij 58), and Triton X-114 in decane. DNA polymerases were not active in AOT, CTAB, and SDS reverse microemulsions, but these enzymes catalyzed DNA synthesis in Brij 58 a nd its mixture with other surfactants. We have also found the system c omposed from the Triton X-114, SDS, CTAB, and Brij 58 (concentration o f 128, 25, 15, and 10 mM, respectively) in hexanol-decane (1:12 v/v), in which DNA polymerases revealed maximum activity. The above system w as optically transparent, fluid, and stable during a few hours with a water-surfactants molar ratio up to 160. The pH dependence of DNA poly merase activity was not significantly different in comparison with wat er; however, DNA polymerase was sensitive to ionic strength in microem ulsions. The dependence of DNA polymerase activity on w(0) was the cur ve with a few optima. DNA polymerases synthesized more products in wat er than in reverse microemulsions, and the processivity of Klenow frag ment decreased. An increase of the water content resulted in an increa se of DNA polymerase processivity. (C) 1998 Elsevier Science B.V. All rights reserved.