STRUCTURAL STUDIES OF BACTERIOPHAGE N4 RNA POLYMERASE AND DNA COMPLEX

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This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. We are using bacteriophage N4 RNA polymerase (N4 RNAP) as a model system to understand transcription mechanism. We have solved apo enzyme structure at 2.3A resolution by using SeMet-protein MAD method and dataset obtained at X25/NSLS. Recently, we have solved N4 RNAP-DNA complex structure by molecular replacement. The dataset was collected with home source X-ray and the resolution is 2.4A. Nobody solved the structure of RNAP-DNA-substrate complex at transcription initiation stage (transcription initiation complex). We prepared initiation complex crystal by co-crystallization and found it diffracts at 2.4A resolution with home source X-ray. To understand molecular detail of transcription imitation reaction, we want to request X25 beamline time to collect transcription initiation complex crystal dataset at atomic resolution. The structure will be the first snap shot of transcription initiation stage and it tells us why RNAP is able to carry out de novo reaction. This project is funded by NIH.