Complete genome sequence of Mesorhizobium ciceri strain R30, a Rhizobium used as a commercial inoculant for Chickpea in Argentina

We report the complete genome sequence of Mesorhizobium ciceri strain R30, a rhizobium strain recommended and used as a commercial inoculant for chickpea in Argentina. The genome consists of almost 7 Mb, distributed into two circular replicons: a chromosome of 6.49 Mb and a plasmid of 0.46 Mb.This work was supported by grants from the Consejo Nacional de Investigaciones Científicas y Técnicas (CONICET), Agencia Nacional de Promoción Científica y Tecnológica (ANPCyT) (PID2020-113207GBI00 funded by MCIN/AEI/10.13039/501100011033); by “ERDF: A Way of Making Europe” (P20_0047), funded by the Junta de Andalucía PAIDI/FEDER/EU; and by the Biotechnology and Biosciences Research Council (BBSRC). We are grateful to Plateforme de Microbiologie Mutualisée (P2M) and the Pasteur International Bioresources network (PIBnet) and to Institut Pasteur Paris for providing the resources for Illumina sequencing. We thank O.G.C. at the Wellcome Centre for Human Genetics for the sequencing data and B.M.R.C. for processing (supported by Wellcome Trust Core Award grant 203141/Z/16/Z and the NIHR Oxford BRC). We are also grateful to Vincent Enouf from Unité de Génétique Moléculaire des Virus à ARN-UMR3569 CNRS, Université de Paris, Centre National de Référence Virus des Infections Respiratoires (dont la grippe) and to F. Sgarlatta for proofreading the manuscript

Biological and structural characterization of theMycobacterium smegmatis nitroreductase NfnB, and its rolein benzothiazinone resistance

Tuberculosis is still a leading cause of death in developing countries, for which there is an urgent need for new pharmacological agents. The synthesis of the novel antimycobacterial drug class of benzothiazinones (BTZs) and the identification of their cellular target as DprE1 (Rv3790), a component of the decaprenylphosphoryl-b-D-ribose 2'-epimerase complex, have been reported recently. Here, we describe the identification and characterization of a novel resistance mechanism to BTZ in Mycobacterium smegmatis. The overexpression of the nitroreductase NfnB leads to the inactivation of the drug by reduction of a critical nitro-group to an amino-group. The direct involvement of NfnB in the inactivation of the lead compound BTZ043 was demonstrated by enzymology, microbiological assays and gene knockout experiments. We also report the crystal structure of NfnB in complex with the essential cofactor flavin mononucleotide, and show that a common amino acid stretch between NfnB and DprE1 is likely to be essential for the interaction with BTZ. We performed docking analysis of NfnB-BTZ in order to understand their interaction and the mechanism of nitroreduction. Although Mycobacterium tuberculosis seems to lack nitroreductases able to inactivate these drugs, our findings are valuable for the design of new BTZ molecules, which may be more effective in vivo

A streamlined, automated protocol for the production of milligram quantities of untagged recombinant rat lactate dehydrogenase A using ÄKTAxpress™

We developed an efficient, automated 2-step purification protocol for the production of milligram quantities of untagged recombinant rat lactate dehydrogenase A (rLDHA) from E. coli, using the ÄKTAxpress™ chromatography system. Cation exchange followed by size exclusion results in average final purity in excess of 93% and yields ~ 14 milligrams per 50 ml of original cell culture in EnPresso B media, in under 8 hrs, including all primary sample processing and column equilibration steps. The protein is highly active and coherent biophysically and a viable alternative to the more problematic human homolog for structural and ligand-binding studies; an apo structure of untagged rLDHA was solved to a resolution 2.29 Å (PDB ID 5ES3). Our automated methodology uses generic commercially available pre-packed columns and simple buffers, and represents a robust standard method for the production of milligram amounts of untagged rLDHA, facilitating a novel fragment screening approach for new inhibitors

Towards new tuberculosis drugs

The need for better drugs to treat tuberculosis has never been greater. Despite insufficient funding for discovery research, intensive efforts have been made to find and develop new lead compounds capable of reducing the duration of the present treatment known as DOTS (directly observed therapy short course), from 6 to under 4 months. This minireview describes the progress achieved during the last 5 years and highlights some of the successes without neglecting the problems