Subunit specialization in AAA+ proteins and substrate unfolding during transcription complex remodeling
Author(s)
Gao, Forson
Ye, Fuzhou
Buck, Martin
Zhang, Xiaodong
Type
Journal Article
Abstract
Bacterial RNA polymerase (RNAP) is a multisubunit enzyme that copies DNA into RNA in a process known as transcription. Bacteria use σ factors to recruit RNAP to promoter regions of genes that need to be transcribed, with 60% bacteria containing at least one specialized σ factor, σ54. σ54 recruits RNAP to promoters of genes associated with stress responses and forms a stable closed complex that does not spontaneously isomerize to the open state where promoter DNA is melted out and competent for transcription. The σ54-mediated open complex formation requires specific AAA+ proteins (ATPases Associated with diverse cellular Activities) known as bacterial enhancer-binding proteins (bEBPs). We have now obtained structures of new intermediate states of bEBP-bound complexes during transcription initiation, which elucidate the mechanism of DNA melting driven by ATPase activity of bEBPs and suggest a mechanistic model that couples the Adenosine triphosphate (ATP) hydrolysis cycle within the bEBP hexamer with σ54 unfolding. Our data reveal that bEBP forms a nonplanar hexamer with the hydrolysis-ready subunit located at the furthest/highest point of the spiral hexamer relative to the RNAP. ATP hydrolysis induces conformational changes in bEBP that drives a vectoral transiting of the regulatory N terminus of σ54 into the bEBP hexamer central pore causing the partial unfolding of σ54, while forming specific bEBP contacts with promoter DNA. Furthermore, our data suggest a mechanism of the bEBP AAA+ protein that is distinct from the hand-over-hand mechanism proposed for many other AAA+ proteins, highlighting the versatile mechanisms utilized by the large protein family.
Date Issued
2025-04-29
Date Acceptance
2025-03-12
Citation
Proceedings of the National Academy of Sciences, 2025, 122 (17)
ISSN
0027-8424
Publisher
Proceedings of the National Academy of Sciences
Journal / Book Title
Proceedings of the National Academy of Sciences
Volume
122
Issue
17
Copyright Statement
© 2025 the Author(s). Published by PNAS. This open access article is distributed under Creative Commons Attribution License 4.0 (CC BY).
License URL
Identifier
10.1073/pnas.2425868122
Subjects
DNA-Directed RNA Polymerases
Bacterial Proteins
DNA-Binding Proteins
Protein Subunits
Adenosine Triphosphate
Transcription, Genetic
Models, Molecular
RNA Polymerase Sigma 54
Promoter Regions, Genetic
ATPases Associated with Diverse Cellular Activities
Publication Status
Published
Article Number
e2425868122
Date Publish Online
2025-04-24