AtfA, a new factor in global regulation of transcription in Acinetobacter spp.

Acinetobacter species are widely distributed bacteria in the environment, and have recently gained notoriety as opportunistic nosocomial pathogens. Here we characterize a novel RNA polymerase-interacting protein named acidic transcription factor A, AtfA. It is small and highly acidic, and is widely distributed throughout the γ proteobacteria, including other significant pathogens in the genera Moraxella, Pseudomonas, Legionella and Vibrio.

Stage-specific fluorescence intensity of GFP and mCherry during sporulation In Bacillus Subtilis.

Background: Fluorescent proteins are powerful molecular biology tools that have been used to study the subcellular dynamics of proteins within live cells for well over a decade. Two fluorescent proteins commonly used to enable dual protein labelling are GFP (green) and mCherry (red). Sporulation in the Gram positive bacterium Bacillus subtilis has been studied for many years as a paradigm for understanding the molecular basis for differential gene expression.

Small subunits of RNA polymerase: localization, levels and implications for core enzyme composition.

Bacterial RNA polymerases (RNAPs) contain several small auxiliary subunits known to co-purify with the core α, β and β' subunits. The ω subunit is conserved between Gram-positive and Gram-negative bacteria, while the δ subunit is conserved within, but restricted to, Gram-positive bacteria. Although various functions have been assigned to these subunits via in vitro assays, very little is known about their in vivo roles.

The structure of bacterial RNA polymerase in complex with the essential transcription elongation factor NusA.

There are three stages of transcribing DNA into RNA. These stages are initiation, elongation and termination, and they are well-understood biochemically. However, despite the plethora of structural information made available on RNA polymerase in the last decade, little is available for RNA polymerase in complex with transcription elongation factors.

Tandem affinity purification vectors for use in gram positive bacteria.

Tandem affinity purification has become a valuable tool for the isolation of protein complexes. Here we describe the construction and use of a series of plasmid vectors for Gram positive bacteria. The vectors utilize the SPA tag as well as variants containing a 3C rather than the TEV protease site as 3C protease has been shown to work efficiently at the low temperatures (4 degrees C) used to isolate protein complexes.

Subcellular partitioning of transcription factors in Bacillus subtilis.

RNA polymerase (RNAP) requires the interaction of various transcription elongation factors to efficiently transcribe RNA. During transcription of rRNA operons, RNAP forms highly processive antitermination complexes by interacting with NusA, NusB, NusG, NusE, and possibly several unidentified factors to increase elongation rates to around twice those observed for mRNA. In previous work we used cytological assays with Bacillus subtilis to identify the major sites of rRNA synthesis within the cell, which are called transcription foci.