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Sleeping sickness is a fatal disease caused by the protozoan parasite Trypanosoma brucei (Tb). Inosine-5'-monophosphate dehydrogenase (IMPDH) has been proposed as a potential drug target, since it maintains the balance between guanylate deoxynucleotide and ribonucleotide levels that is pivotal for the parasite. Here we report the structure of TbIMPDH at room temperature utilizing free-electron laser radiation on crystals grown in living insect cells. The 2.80 Å resolution structure reveals the presence of ATP and GMP at the canonical sites of the Bateman domains, the latter in a so far unknown coordination mode. Consistent with previously reported IMPDH complexes harboring guanosine nucleotides at the second canonical site, TbIMPDH forms a compact oligomer structure, supporting a nucleotide-controlled conformational switch that allosterically modulates the catalytic activity. The oligomeric TbIMPDH structure we present here reveals the potential of in cellulo crystallization to identify genuine allosteric co-factors from a natural reservoir of specific compounds.

Trypanosoma brucei inosine-5′-monophosphate dehydrogenase (IMPDH) is an enzyme in the guanine nucleotide biosynthesis pathway and of interest as a drug target. Here the authors present the 2.8 Å room temperature structure of TbIMPDH determined by utilizing X-ray free-electron laser radiation and crystals that were grown in insect cells and find that ATP and GMP are bound at the canonical sites of the Bateman domains.

X-ray crystallography requires sufficiently large crystals to obtain structural insights at atomic resolution, routinely obtained in vitro by time-consuming screening. Recently, successful data collection was reported from protein microcrystals grown within living cells using highly brilliant free-electron laser and third-generation synchrotron radiation. Here, we analyzed in vivo crystal growth of firefly luciferase and Green Fluorescent Protein-tagged reovirus μNS by live-cell imaging, showing that dimensions of living cells did not limit crystal size. The crystallization process is highly dynamic and occurs in different cellular compartments. In vivo protein crystallization offers exciting new possibilities for proteins that do not form crystals in vitro.

To assess the impact of the implementation of a Computerized Physician Order Entry (CPOE) associated with a pharmaceutical checking of medication orders on medication errors in the 3 stages of drug management (i.e. prescription, dispensing and administration) in an orthopaedic surgery unit.A before-after observational study was conducted in the 66-bed orthopaedic surgery unit of a teaching hospital (700 beds) in Paris France. Direct disguised observation was used to detect errors in prescription, dispensing and administration of drugs, before and after the introduction of computerized prescriptions. Compliance between dispensing and administration on the one hand and the medical prescription on the other hand was studied. The frequencies and types of errors in prescribing, dispensing and administration were investigated.During the pre and post-CPOE period (two days for each period) 111 and 86 patients were observed, respectively, with corresponding 1,593 and 1,388 prescribed drugs. The use of electronic prescribing led to a significant 92% decrease in prescribing errors (479/1593 prescribed drugs (30.1%) vs 33/1388 (2.4%), p < 0.0001) and to a 17.5% significant decrease in administration errors (209/1222 opportunities (17.1%) vs 200/1413 (14.2%), p < 0.05). No significant difference was found in regards to dispensing errors (430/1219 opportunities (35.3%) vs 449/1407 (31.9%), p = 0.07).The use of CPOE and a pharmacist checking medication orders in an orthopaedic surgery unit reduced the incidence of medication errors in the prescribing and administration stages. The study results suggest that CPOE is a convenient system for improving the quality and safety of drug management.