Dissection of the β subunit in the Escherichia coli RNA polymerase into domains by proteolytic cleavage

The 1342 amino acid long β subunit of Escherichia coli RNA polymerase includes a dispensable region (residues 940-1040) that is absent in homologous RNA polymerase subunits from chloroplasts, eukaryotes, and archaebacteria (Borukhov, S., Severinov, K., Kashlev, M., Lebedev, A., Bass, I., Rowland, G. C., Lim, P.-P., Glass, R. E., Nikiforov, V., and Goldfarb, A. (1991) J. Biol. Chem. 266, 23921-23926). Genetic disruption of this region by in-frame deletion or insertion sensitizes the β subunit in assembled RNA polymerase molecules to attack by trypsin. We demonstrate that RNA polymerase with the β polypeptide cleaved in the dispensable region retains normal in vitro activity. Moreover, the RNA polymerase activity is completely restored after denaturation and reconstitution of the enzyme carrying cleaved β subunit indicating that its carboxyl- and amino-terminal parts fold and assemble into RNA polymerase as separate entities.