title: Structure and function of the mycobacterial transcription initiation complex with the essential regulator RbpA.
abstract: RbpA and CarD are essential transcription regulators in mycobacteria. Mechanistic analyses of promoter open complex (RPo) formation establish that RbpA and CarD cooperatively stimulate formation of an intermediate (RP2) leading to RPo; formation of RP2 is likely a bottleneck step at the majority of mycobacterial promoters. Once RPo forms, CarD also disfavors its isomerization back to RP2. We determined a 2.76 Å-resolution crystal structure of a mycobacterial transcription initiation complex (TIC) with RbpA as well as a CarD/RbpA/TIC model. Both CarD and RbpA bind near the upstream edge of the -10 element where they likely facilitate DNA bending and impede transcription bubble collapse. In vivo studies demonstrate the essential role of RbpA, show the effects of RbpA truncations on transcription and cell physiology, and indicate additional functions for RbpA not evident in vitro. This work provides a framework to understand the control of mycobacterial transcription by RbpA and CarD.
authors.affiliation: The Rockefeller University, New York, United States.
authors.forename: Elizabeth A
authors.initials: EA
authors.lastname: Hubin
authors.affiliation: Immunology Program, Sloan-Kettering Institute, New York, United States.
authors.forename: Allison
authors.initials: A
authors.lastname: Fay
authors.affiliation: The Rockefeller University, New York, United States.
authors.forename: Catherine
authors.initials: C
authors.lastname: Xu
authors.affiliation: Immunology Program, Sloan-Kettering Institute, New York, United States.
authors.forename: James M
authors.initials: JM
authors.lastname: Bean
authors.affiliation: Independent Researcher, Madison, United States.
authors.forename: Ruth M
authors.initials: RM
authors.lastname: Saecker
authors.affiliation: Immunology Program, Sloan-Kettering Institute, New York, United States.|Division of Infectious Diseases, Memorial Sloan-Kettering Cancer Center, New York, United States.
authors.forename: Michael S
authors.identifier: 0000-0001-7918-5164
authors.initials: MS
authors.lastname: Glickman
authors.affiliation: The Rockefeller University, New York, United States.
authors.forename: Seth A
authors.identifier: 0000-0002-8241-3153
authors.initials: SA
authors.lastname: Darst
authors.affiliation: The Rockefeller University, New York, United States.
authors.forename: Elizabeth A
authors.initials: EA
authors.lastname: Campbell
mesh_terms: D001426:Bacterial Proteins / Q000737:chemistry* / Q000378:metabolism*; D018360:Crystallography, X-Ray; D008958:Models, Molecular; D009097:Multienzyme Complexes / Q000737:chemistry / Q000378:metabolism; D009161:Mycobacterium / Q000201:enzymology* / Q000235:genetics / Q000378:metabolism*; D011401:Promoter Regions, Genetic; D011487:Protein Conformation; D014157:Transcription Factors / Q000737:chemistry* / Q000378:metabolism*; D061785:Transcription Initiation, Genetic
chemical_list: D001426:Bacterial Proteins; D009097:Multienzyme Complexes; D014157:Transcription Factors
keywords: E. coli; M. bovis; M. smegmatis; M. tuberculosis; biochemistry; biophysics; structural biology