Process for reconstituting the polymerase III* and other subassemblies of E. coli DNA polymerase III holoenzyme from peptide subunits

摘要

The process of the invention provides for the reconstitution of the polymerase III* subassembly, Pol III*, of E. coli DNA polymerase III holoenzyme from substantially pure peptide subunits. In the first of two general schemes in which the subunits are added in a specified order, gamma and tau are premixed before addition of delta and delta '. In the second general scheme, delta ' is first assembled onto gamma (or tau ); then the excess delta ' is removed before adding tau (or gamma ), following which delta is added. Reconstituted Pol III* had the same subunit composition as purified natural Pol III*, as well as similar activity. Other smaller subassemblies of the polymerase III holoenzyme may also be reconstituted by the process of the invention.