The crystal structure of the mycobacterial DprE1 reveals how the TB drug benzothiazinone BTZ043 blocks this microbial enzyme target.

Burkholderia cenocepacia is an opportunistic pathogen particularly dangerous for cystic fibrosis (CF) patients. It can cause a severe decline in CF lung function possibly developing into a life-threatening systemic infection known as cepacia syndrome. Antibiotic resistance and presence of numerous virulence determinants in the genome make B. cenocepacia extremely difficult to treat. Better understanding of its resistance profiles and mechanisms is crucial to improve management of these infections. Here we present the clinical distribution of B. cenocepacia described in the last six years and methods for identification and classification of epidemic strains. We also detail new antibiotics, clinical trials and alternative approaches reported in the literature in the last five years to tackle B. cenocepacia resistance issue. All together these findings point out the urgent need of new and alternative therapies to improve CF patients’ life expectancy.

Pyruvate kinase (PK) is critical for the regulation of the glycolytic pathway. The regulatory properties ofEscherichia coli were investigated by mutating six charged residues involved in interdomain salt bridges (Arg²⁷¹, Arg²⁹², Asp²⁹⁷, and Lys⁴¹³) and in the binding of the allosteric activator (Lys³⁸² and Arg⁴³¹). Arg²⁷¹ and Lys⁴¹³ are located at the interface between A and C domains within one subunit. The R271L and K413Q mutant enzymes exhibit altered kinetic properties. In K413Q, there is partial enzyme activation, whereas R271L is characterized by a bias toward the T-state in the allosteric equilibrium. In the T-state, Arg²⁹² and Asp²⁹⁷form an intersubunit salt bridge. The mutants R292D and D297R are totally inactive. The crystal structure of R292D reveals that the mutant enzyme retains the T-state quaternary structure. However, the mutation induces a reorganization of the interface with the creation of a network of interactions similar to that observed in the crystal structures of R-state yeast and M1 PK proteins. Furthermore, in the R292D structure, two loops that are part of the active site are disordered. The K382Q and R431E mutations were designed to probe the binding site for fructose 1,6-bisphosphate, the allosteric activator. R431E exhibits only slight changes in the regulatory properties. Conversely, K382Q displays a highly altered responsiveness to the activator, suggesting that Lys³⁸² is involved in both activator binding and allosteric transition mechanism. Taken together, these results support the notion that domain interfaces are critical for the allosteric transition. They couple changes in the tertiary and quaternary structures to alterations in the geometry of the fructose 1,6-bisphosphate and substrate binding sites. These site-directed mutagenesis data are discussed in the light of the molecular basis for the hereditary nonspherocytic hemolytic anemia, which is caused by mutations in human erythrocyte PK gene.

Nontuberculous mycobacteria (NTM) have recently emerged as important pathogens among cystic fibrosis (CF) patients worldwide. Mycobacterium abscessus is becoming the most worrisome NTM in this cohort of patients and recent findings clarified why this pathogen is so prone to this disease. M. abscessus drug therapy takes up to 2 years and its failure causes an accelerated lung function decline. The M. abscessus colonization of lung alveoli begins with smooth strains producing glycopeptidolipids and biofilm, whilst in the invasive infection, “rough” mutants are responsible for the production of trehalose dimycolate, and consequently, cording formation. Human-to-human M. abscessus transmission was demonstrated among geographically separated CF patients by whole-genome sequencing of clinical isolates worldwide. Using a M. abscessus infected CF zebrafish model, it was demonstrated that CFTR (cystic fibrosis transmembrane conductance regulator) dysfunction seems to have a specific role in the immune control of M. abscessus infections only. This pathogen is also intrinsically resistant to many drugs, thanks to its physiology and to the acquisition of new mechanisms of drug resistance. Few new compounds or drug formulations active against M. abscessus are present in preclinical and clinical development, but recently alternative strategies have been investigated, such as phage therapy and the use of β-lactamase inhibitors.