Normal-mode analysis suggests protein flexibility modulation throughout RNA polymerase’s functional cycle. Biochemistry 43, 13083–13096.Wilson

ABSTRACT: To explore the domain-scale flexibility of bacterial RNA polymerase (RNAP) throughout its functional cycle, block normal-mode analyses (BNM) were performed on several important functional states, including the holoenzyme, the core complex, a model of RNAP bound to primarily duplex DNA, and a model of the ternary elongation complex. The calculations utilized a molecular mechanics (MM) force field with physical interactions; this is made possible by the use of BNM and the implementation of a sparse-matrix diagonalization routine. The use of homology models necessitated the MM force field rather than the simpler elastic network model (ENM). From the MM/BNM, we have systematically and semiquantitatively calculated the atomic fluctuations in the four functional states without bias due to crystal packing or other artifactual forces. We have observed that both R subunits and the ö subunit are rigid, in line with their roles as structural motifs that are not mechanistically involved in RNAP’s functional cycle. It has been observed that the â subunit has two highly mobile domains; these are commonly known as the â1 and â2 domains. Our calculations suggest that the flexibility of these domains is modulated throughout the functional cycle and that they move entirely independently of each other unless DNA is bound. From an energetic perspective, we have shown the â2 domain can flex into and out of the cleft, forming interactions with DNA in the TEC as has been previously proposed. Our calculations also confir