An in vivo crosslinking system for identifying mycobacterial protein-protein interactions.

The analysis of protein-protein interactions in Mycobacterium tuberculosis has the potential to shed light on the functions of the large number of predicted open-reading frames annotated as conserved hypothetical proteins. We have developed a formaldehyde crosslinking system to detect in vivo interactions in mycobacteria. Our Gateway-adapted vector system uses three promoter strengths, including constitutive and regulatable versions, for the expression of target proteins with either an N- or C-terminal His-Strep-Strep tag.

The mucoid switch in Pseudomonas aeruginosa represses quorum sensing systems and leads to complex changes to stationary phase virulence factor regulation.

The opportunistic pathogen Pseudomonas aeruginosa chronically infects the airways of Cystic Fibrosis (CF) patients during which it adapts and undergoes clonal expansion within the lung. It commonly acquires inactivating mutations of the anti-sigma factor MucA leading to a mucoid phenotype, caused by excessive production of the extracellular polysaccharide alginate that is associated with a decline in lung function. Alginate production is believed to be the key benefit of mucA mutations to the bacterium in the CF lung.

Induced ectopic expression of HigB toxin in Mycobacterium tuberculosis results in growth inhibition, reduced abundance of a subset of mRNAs and cleavage of tmRNA.

In Mycobacterium tuberculosis, the genes Rv1954A-Rv1957 form an operon that includes Rv1955 and Rv1956 which encode the HigB toxin and the HigA antitoxin respectively. We are interested in the role and regulation of this operon, since toxin-antitoxin systems have been suggested to play a part in the formation of persister cells in mycobacteria. To investigate the function of the higBA locus, effects of toxin expression on mycobacterial growth and transcript levels were assessed in M.

Ethnicity and mycobacterial lineage as determinants of tuberculosis disease phenotype.

Background: Emerging evidence suggests that Mycobacterium tuberculosis (Mtb) lineage and host ethnicity can determine tuberculosis (TB) clinical disease patterns but their relative importance and interaction are unknown.

Methods: We evaluated prospectively collected TB surveillance and Mtb strain typing data in an ethnically heterogeneous UK population. Lineage assignment was denoted using 15-loci mycobacterial interspersed repetitive units containing variable numbers of tandem repeats (MIRU-VNTR) and MIRU-VNTRplus.

Substituted aminopyrimidine protein kinase B (PknB) inhibitors show activity against Mycobacterium tuberculosis.

A high-throughput screen against PknB, an essential serine-threonine protein kinase present in Mycobacterium tuberculosis (M. tuberculosis), allowed the identification of an aminoquinazoline inhibitor which was used as a starting point for SAR investigations. Although a significant improvement in enzyme affinity was achieved, the aminoquinazolines showed little or no cellular activity against M.

Effective inhibitors of the essential kinase PknB and their potential as anti-mycobacterial agents.

PknB is an essential serine/threonine kinase of Mycobacterium tuberculosis with possible roles in a number of signalling pathways involved in cell division and metabolism. We screened a library of >50,000 compounds for inhibitors of the in vitro phosphorylation of GarA (Rv1827) by PknB and identified a number of inhibitors. A program of synthetic medicinal chemistry was subsequently conducted around one class of inhibitors and was successful in generating ATP competitive inhibitors with potency in the nanomolar range.

New anti-tuberculosis agents amongst known drugs.

Mycobacterium tuberculosis has an on-going impact on global public health and new therapeutics to treat tuberculosis are urgently required. The emergence of drug resistant tuberculosis poses a serious threat to the control of this pathogen, and the development of drugs that are active against the resistant strains is vital. A medium-throughput assay using the Alamar Blue reagent was set-up to identify novel inhibitors of M.