7-(5-Methyl­sulfanyl-β-d-erythrofuran­osyl)-7H-pyrrolo­[2,3-d]pyrimidin-4-amine monohydrate (MT-tubercidin·H2O)

The title compound, C12H16N4O3S·H2O, which has potential as a possible anti­malarial drug, was studied when small deviations in melting points, for two differently aged preparations, were observed. The unexpected existence of a water mol­ecule of crystallization is considered to be the cause of this variation. The 7H-pyrrolo­[2,3-d]pyrimidine unit is very slightly puckered with a total puckering amplitude of 0.035 (2) Å; its mean plane makes an angle of 88.40 (12)° with the mean plane through the ribofuranosyl unit. In the crystal, the mol­ecules are bound by strong O—H⋯N and N—H⋯O hydrogen bonds, utilizing all available protons and linking mainly through the water of crystallization

Validation of Plasmodium falciparum dUTPase as the target of 5'-tritylated deoxyuridine analogues with anti-malarial activity

BACKGROUND: Malaria remains as a major global problem, being one of the infectious diseases that engender highest mortality across the world. Due to the appearance of resistance and the lack of an effective vaccine, the search of novel anti-malarials is required. Deoxyuridine 5'-triphosphate nucleotido-hydrolase (dUTPase) is responsible for the hydrolysis of dUTP to dUMP within the parasite and has been proposed as an essential step in pyrimidine metabolism by providing dUMP for thymidylate biosynthesis. In this work, efforts to validate dUTPase as a drug target in Plasmodium falciparum are reported. METHODS: To investigate the role of PfdUTPase in cell survival different strategies to generate knockout mutants were used. For validation of PfdUTPase as the intracellular target of four inhibitors of the enzyme, mutants overexpressing PfdUTPase and HsdUTPase were created and the IC50 for each cell line with each compound was determined. The effect of these compounds on dUTP and dTTP levels from P. falciparum was measured using a DNA polymerase assay. Detailed localization studies by indirect immunofluorescence microscopy and live cell imaging were also performed using a cell line overexpressing a Pfdut-GFP fusion protein. RESULTS:Different attempts of disruption of the dut gene of P. falciparum were unsuccessful while a 3' replacement construct could recombine correctly in the locus suggesting that the enzyme is essential. The four 5'-tritylated deoxyuridine analogues described are potent inhibitors of the P. falciparum dUTPase and exhibit antiplasmodial activity. Overexpression of the Plasmodium and human enzymes conferred resistance against selective compounds, providing chemical validation of the target and confirming that indeed dUTPase inhibition is involved in anti-malarial activity. In addition, incubation with these inhibitors was associated with a depletion of the dTTP pool corroborating the central role of dUTPase in dTTP synthesis. PfdUTPase is mainly localized in the cytosol. CONCLUSION: These results strongly confirm the pivotal and essential role of dUTPase in pyrimidine biosynthesis of P. falciparum intraerythrocytic stages

The Judge and His Clerks

In his memoir, Life and Times in the Three Branches, Judge Coffin recounts the history of the institution of the law clerk and observes, “I was the first such creature Maine had seen.” He served as a clerk from April 1947 to June 1949 for United States District Court Judge John D. Clifford, working in the same chambers that he would later inhabit as a judge. Over the course of his more than four decades on the Court of Appeals, Judge Coffin would have sixty-eight clerks of his own. Those of us lucky enough to be in that family of sixty-eight had an incomparable experience both personally and professionally. One of our number, Peter Byrne, captured the essence of a Coffin clerkship in an essay that was published in this journal in 1991. When asked about including a description of clerking for Judge Coffin in this tribute issue, we immediately thought of Peter’s piece, which remains a perfect reflection of the Coffin clerkship experience and the special relationship we each had with him

Anionic lipids unlock the gates of select ion channels in the pacemaker family

Lipids play important roles in regulating membrane protein function, but the molecular mechanisms used are elusive. Here we investigated how anionic lipids modulate SthK, a bacterial pacemaker channel homolog, and HCN2, whose activity contributes to pacemaking in the heart and brain. Using SthK allowed the reconstitution of purified channels in controlled lipid compositions for functional and structural assays that are not available for the eukaryotic channels. We identified anionic lipids bound tightly to SthK and their exact binding locations and determined that they potentiate channel activity. Cryo-EM structures in the most potentiating lipids revealed an open state and identified a nonannular lipid bound with its headgroup near an intersubunit salt bridge that clamps the intracellular channel gate shut. Breaking this conserved salt bridge abolished lipid modulation in SthK and eukaryotic HCN2 channels, indicating that anionic membrane lipids facilitate channel opening by destabilizing these interactions. Our findings underline the importance of state-dependent protein-lipid interactions

Development of Multiple Job Performance Measures in a Representative Sample of Jobs

The goal of criterion development in Project A was to construct multiple measures of the major components of job performance such that the total performance domain for a representative sample of the population of entry‐level enlisted positions in the U.S. Army was covered. These measures were to be used as criteria against which to validate both experimental and existing predictors of job performance. The initial model specified that performance is multidimensional within two major categories of dimensions designated as organization‐wide and job specific. The development strategy involved describing the total domain of job content via extensive task analyses and critical incident analyses, generating the critical performance dimensions that constitute it, constructing measures for each dimension, and evaluating each measure using expert judgment and field test data. The specific measures developed consisted of rating scales, tests of job knowledge, hands‐on job samples, and archival records. The major steps in the job analyses, content sampling, instrument construction, and instrument evaluation are described, and the final array of criterion measures is presented