RNA polymerase assembly in vitro. Temperature dependence of reactivation of denatured core enzyme.

The Escherichia coli RNA polymerase core molecule, after denaturation in 6 M guanidine hydrochloride, can be completely reactivated in the absence of sigma subunit. Reactivation is temperature dependent. At 4 degrees a renatured-inactive preparation is formed that has most of the secondary structure of the original native molecule but has a reduced sedimentation coefficient and a smaller Stokes radius and is, therefore, of lower molecular weight. Upon warming to 37 degrees the renatured-inactive preparation is converted in a time-dependent process to the renatured-active preparation, which has the same amount of secondary structure and same molecular weight as native RNA polymerase. Since the renatured-inactive material is probably composed of subunit assemblies and can be readily reactivated, it should be useful for studying the subunit interactions and control of assembly of RNA polymerase.