Molecular insights into RNA polymerase recycling by mycobacterial HelD

Mycobacterial HelD is a transcription factor that binds RNA polymerase (RNAP) and thereby rescues it from stalled transcription complexes. HelD also protects RNAP against the antibiotic rifampicin. HelD, however, must be released from the rescued RNAP to participate in the next round of the transcription cycle. The exact mechanism of this release is unknown. Here we show that HelD from Mycobacterium smegmatis can form a complex with RNAP, together with σA primary sigma factor, and RbpA, but not CarD, transcription factors. Using cryo-EM, we solved a series of RNAP-σA-RbpA-HelD structures with or without promoter DNA. These snapshots capture the involvement of HelD in transcription initiation, describes mechanistic aspects of HelD release, and its protective effect against rifampicin. Biochemical evidence supports these findings, defines the role of ATP binding/hydrolysis to/by HelD in the process, and functionally confirms the rifampicin-protective effect of HelD. Taken together, HelD assists an alternative pathway of transcription initiation.