Structure and function of the Rad9-binding region of the DNA-damage checkpoint adaptor TopBP1

TopBP1 is a scaffold protein that coordinates activation of the DNA-damage-checkpoint response by coupling binding of the 9-1-1 checkpoint clamp at sites of ssDNA, to activation of the ATR-ATRIP checkpoint kinase complex. We have now determined the crystal structure of the N-terminal region of human TopBP1, revealing an unexpected triple-BRCT domain structure. The arrangement of the BRCT domains differs significantly from previously described tandem BRCT domain structures, and presents two distinct sites for binding phosphopeptides in the second and third BRCT domains. We show that the site in the second but not third BRCT domain in the N-terminus of TopBP1, provides specific interaction with a phosphorylated motif at pSer387 in Rad9, which can be generated by CK2

Structure of fructose bisphosphate aldolase from Bartonella henselae bound to fructose 1,6-bisphosphate

While other aldolases crystallize readily in the apo form, diffraction-quality crystals of B. henselae aldolase could only be obtained in the presence of the native substrate. The quaternary structure is tetrameric, as is typical of aldolases

Subjectivation and performative politics—Butler thinking Althusser and Foucault: intelligibility, agency and the raced-nationed-religioned subjects of education

Judith Butler is perhaps best known for her take-up of the debate between Derrida and Austin over the function of the performative and her subsequent suggestion that the subject be understood as performatively constituted. Another important but less often noted move within Butler‘s consideration of the processes through which the subject is constituted is her thinking between Althusser‘s notion of subjection and Foucault‘s notion of subjectivation. In this paper, I explore Butler‘s understanding of processes of subjectivation; examine the relationship between subjectivation and the performative suggested in and by Butler‘s work, and consider how the performative is implicated in processes of subjectivation – in =who‘ the subject is, or might be, subjectivated as. Finally, I examine the usefulness of understanding the subjectivating effects of discourse for education, in particular for educationalists concerned to make better sense of and interrupt educational inequalities. In doing this I offer a reading of an episode of ethnographic data generated in an Australian high School. I suggest that it is through subjectivating processes of the sort that Butler helps us to understand that some students are rendered subjects inside the educational endeavour, and others are rendered outside this endeavour or, indeed, outside student-hood

Structure of fructose bisphosphate aldolase from Encephalitozoon cuniculi

The eukaryotic parasite E. cuniculi expresses a fructose bisphosphate aldolase that crystallizes readily in the presence of the partial substrate analog phosphate. This aldolase–phosphate structure and that of the sugar-bound Schiff base are reported. E. cuniculi aldolase displays a dimeric structure rather than the expected tetrameric quaternary structure

The statistical neuroanatomy of frontal networks in the macaque

We were interested in gaining insight into the functional properties of frontal networks based upon their anatomical inputs. We took a neuroinformatics approach, carrying out maximum likelihood hierarchical cluster analysis on 25 frontal cortical areas based upon their anatomical connections, with 68 input areas representing exterosensory, chemosensory, motor, limbic, and other frontal inputs. The analysis revealed a set of statistically robust clusters. We used these clusters to divide the frontal areas into 5 groups, including ventral-lateral, ventral-medial, dorsal-medial, dorsal-lateral, and caudal-orbital groups. Each of these groups was defined by a unique set of inputs. This organization provides insight into the differential roles of each group of areas and suggests a gradient by which orbital and ventral-medial areas may be responsible for decision-making processes based on emotion and primary reinforcers, and lateral frontal areas are more involved in integrating affective and rational information into a common framework

The effect of health literacy on knowledge and receipt of colorectal cancer screening: a survey study

BACKGROUND: An estimated one-half of Americans have limited health literacy skills. Low literacy has been associated with less receipt of preventive services, but its impact on colorectal cancer (CRC) screening is unclear. We sought to determine whether low literacy affects patients' knowledge or receipt of CRC screening. METHODS: Pilot survey study of patients aged 50 years and older at a large, university-affiliated internal medicine practice. We assessed patients' knowledge and receipt of CRC screening, basic sociodemographic information, and health literacy level. We defined limited literacy as reading below the ninth grade level as determined by the Rapid Estimate of Adult Literacy in Medicine. Bivariate analyses and exact logistic regression were used to determine the association of limited health literacy with knowledge and receipt of CRC screening. RESULTS: We approached 105 patients to yield our target sample of 50 completing the survey (recruitment rate 48%). Most subjects were female (72%), African-American (58%), and had household incomes less than $25,000 (87%). Overall, 48% of patients had limited literacy skills (95% CI 35% to 61%). Limited literacy patients were less likely than adequate literacy patients to be able to name or describe any CRC screening test (50% vs. 96%, p < 0.01). In the multivariable model, limited literacy patients were 44% less likely to be knowledgeable of CRC screening (RR 0.56, p < 0.01). Self-reported screening rates were similar (54% vs. 58%, p = 0.88). CONCLUSION: Patients with limited literacy skills are less likely to be knowledgeable of CRC screening compared to adequate literacy patients. Primary care providers should ensure patients' understanding of CRC screening when discussing screening options. Further research is needed to determine if educating low literacy patients about CRC screening can increase screening rates

Piperidinols that show anti-tubercular activity as inhibitors of arylamine N-acetyltransferase: an essential enzyme for mycobacterial survival inside macrophages

Latent M. tuberculosis infection presents one of the major obstacles in the global eradication of tuberculosis (TB). Cholesterol plays a critical role in the persistence of M. tuberculosis within the macrophage during latent infection. Catabolism of cholesterol contributes to the pool of propionyl-CoA, a precursor that is incorporated into cell-wall lipids. Arylamine N-acetyltransferase (NAT) is encoded within a gene cluster that is involved in the cholesterol sterol-ring degradation and is essential for intracellular survival. The ability of the NAT from M. tuberculosis (TBNAT) to utilise propionyl-CoA links it to the cholesterol-catabolism pathway. Deleting the nat gene or inhibiting the NAT enzyme prevents intracellular survival and results in depletion of cell-wall lipids. TBNAT has been investigated as a potential target for TB therapies. From a previous high-throughput screen, 3-benzoyl-4-phenyl-1-methylpiperidinol was identified as a selective inhibitor of prokaryotic NAT that exhibited antimycobacterial activity. The compound resulted in time-dependent irreversible inhibition of the NAT activity when tested against NAT from M. marinum (MMNAT). To further evaluate the antimycobacterial activity and the NAT inhibition of this compound, four piperidinol analogues were tested. All five compounds exert potent antimycobacterial activity against M. tuberculosis with MIC values of 2.3-16.9 µM. Treatment of the MMNAT enzyme with this set of inhibitors resulted in an irreversible time-dependent inhibition of NAT activity. Here we investigate the mechanism of NAT inhibition by studying protein-ligand interactions using mass spectrometry in combination with enzyme analysis and structure determination. We propose a covalent mechanism of NAT inhibition that involves the formation of a reactive intermediate and selective cysteine residue modification. These piperidinols present a unique class of antimycobacterial compounds that have a novel mode of action different from known anti-tubercular drugs

Influence of flow rate and scaffold pore size on cell behavior during mechanical stimulation in a flow perfusion bioreactor.

Mechanically stimulating cell-seeded scaffolds by flow-perfusion is one approach utilized for developing clinically applicable bone graft substitutes. A key challenge is determining the magnitude of stimuli to apply that enhances cell differentiation but minimizes cell detachment from the scaffold. In this study, we employed a combined computational modeling and experimental approach to examine how the scaffold mean pore size influences cell attachment morphology and subsequently impacts upon cell deformation and detachment when subjected to fluid-flow. Cell detachment from osteoblast-seeded collagen-GAG scaffolds was evaluated experimentally across a range of scaffold pore sizes subjected to different flow rates and exposure times in a perfusion bioreactor. Cell detachment was found to be proportional to flow rate and inversely proportional to pore size. Using this data, a theoretical model was derived that accurately predicted cell detachment as a function of mean shear stress, mean pore size, and time. Computational modeling of cell deformation in response to fluid flow showed the percentage of cells exceeding a critical threshold of deformation correlated with cell detachment experimentally and the majority of these cells were of a bridging morphology (cells stretched across pores). These findings will help researchers optimize the mean pore size of scaffolds and perfusion bioreactor operating conditions to manage cell detachment when mechanically simulating cells via flow perfusion. Biotechnol. Bioeng. © 2012 Wiley Periodicals, Inc

The structure of sedoheptulose-7-phosphate isomerase from Burkholderia pseudomallei reveals a zinc binding site at the heart of the active site

Copyright © 2010 Elsevier. NOTICE: this is the author’s version of a work that was accepted for publication in Journal of Molecular Biology. Changes resulting from the publishing process, such as peer review, editing, corrections, structural formatting, and other quality control mechanisms may not be reflected in this document. Changes may have been made to this work since it was submitted for publication. A definitive version was subsequently published in Journal of Molecular Biology, 2010, Vol. 400, Issue 3, pp. 379 – 392 DOI: http://dx.doi.org/10.1016/j.jmb.2010.04.058Heptoses are found in the surface polysaccharides of most bacteria, contributing to structures that are essential for virulence and antibiotic resistance. Consequently, the biosynthetic enzymes for these sugars are attractive targets for novel antibiotics. The best characterized biosynthetic enzyme is GmhA, which catalyzes the conversion of sedoheptulose-7-phosphate into D-glycero-D-manno-heptopyranose-7-phosphate, the first step in the biosynthesis of heptose. Here, the structure of GmhA from Burkholderia pseudomallei is reported. This enzyme contains a zinc ion at the heart of its active site: this ion stabilizes the active, closed form of the enzyme and presents coordinating side chains as a potential acid and base to drive catalysis. A complex with the product demonstrates that the enzyme retains activity in the crystal and thus suggests that the closed conformation is catalytically relevant and is an excellent target for the development of therapeutics. A revised mechanism for the action of GmhA is postulated on the basis of this structure and the activity of B. pseudomallei GmhA mutants