Dataset Information

Structural, functional, and genetic analyses of the actinobacterial transcription factor RbpA.

ABSTRACT: Gene expression is highly regulated at the step of transcription initiation, and transcription activators play a critical role in this process. RbpA, an actinobacterial transcription activator that is essential in Mycobacterium tuberculosis (Mtb), binds selectively to group 1 and certain group 2 ?-factors. To delineate the molecular mechanism of RbpA, we show that the Mtb RbpA ?-interacting domain (SID) and basic linker are sufficient for transcription activation. We also present the crystal structure of the Mtb RbpA-SID in complex with domain 2 of the housekeeping ?-factor, ?(A). The structure explains the basis of ?-selectivity by RbpA, showing that RbpA interacts with conserved regions of ?(A) as well as the nonconserved region (NCR), which is present only in housekeeping ?-factors. Thus, the structure is the first, to our knowledge, to show a protein interacting with the NCR of a ?-factor. We confirm the basis of selectivity and the observed interactions using mutagenesis and functional studies. In addition, the structure allows for a model of the RbpA-SID in the context of a transcription initiation complex. Unexpectedly, the structural modeling suggests that RbpA contacts the promoter DNA, and we present in vivo and in vitro studies supporting this finding. Our combined data lead to a better understanding of the mechanism of RbpA function as a transcription activator.

SUBMITTER: Hubin EA

PROVIDER: S-EPMC4466734 | biostudies-literature | 2015 Jun

REPOSITORIES: biostudies-literature

ACCESS DATA

Publications

Structural, functional, and genetic analyses of the actinobacterial transcription factor RbpA.

Hubin Elizabeth A EA Tabib-Salazar Aline A Humphrey Laurence J LJ Flack Joshua E JE Olinares Paul Dominic B PD Darst Seth A SA Campbell Elizabeth A EA Paget Mark S MS

Proceedings of the National Academy of Sciences of the United States of America 20150526 23

Gene expression is highly regulated at the step of transcription initiation, and transcription activators play a critical role in this process. RbpA, an actinobacterial transcription activator that is essential in Mycobacterium tuberculosis (Mtb), binds selectively to group 1 and certain group 2 σ-factors. To delineate the molecular mechanism of RbpA, we show that the Mtb RbpA σ-interacting domain (SID) and basic linker are sufficient for transcription activation. We also present the crystal str ...[more]

PMID: 26040003

Similar Datasets

Project description:BackgroundThe post-genomic era is characterised by a torrent of biological information flooding the public databases. As a direct consequence, similarity searches starting with a single query sequence frequently lead to the identification of hundreds, or even thousands of potential homologues. The huge volume of data renders the subsequent structural, functional and evolutionary analyses very difficult. It is therefore essential to develop new strategies for efficient sampling of this large sequence space, in order to reduce the number of sequences to be processed. At the same time, it is important to retain the most pertinent sequences for structural and functional studies.ResultsAn exhaustive analysis on a large scale test set (284 protein families) was performed to compare the efficiency of four different sampling methods aimed at selecting the most pertinent sequences. These four methods sample the proteins detected by BlastP searches and can be divided into two categories: two customisable methods where the user defines either the maximal number or the percentage of sequences to be selected; two automatic methods in which the number of sequences selected is determined by the program. We focused our analysis on the potential information content of the sampled sets of sequences using multiple alignment of complete sequences as the main validation tool. The study considered two criteria: the total number of sequences in BlastP and their associated E-values. The subsequent analyses investigated the influence of the sampling methods on the E-value distributions, the sequence coverage, the final multiple alignment quality and the active site characterisation at various residue conservation thresholds as a function of these criteria.ConclusionThe comparative analysis of the four sampling methods allows us to propose a suitable sampling strategy that significantly reduces the number of homologous sequences required for alignment, while at the same time maintaining the relevant information concerning the active site residues.

Dataset Information

Structural, functional, and genetic analyses of the actinobacterial transcription factor RbpA.

Publications

Structural, functional, and genetic analyses of the actinobacterial transcription factor RbpA.

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure

Tweets