Inositol 1,3,4,5,6-pentakisphosphate 2-kinase is a distant IPK member with a singular inositide binding site for axial 2-OH recognition

Inositol phosphates (InsPs) are signaling molecules with multiple roles in cells. In particular Graphic (InsP6) is involved in mRNA export and editing or chromatin remodeling among other events. InsP6 accumulates as mixed salts (phytate) in storage tissues of plants and plays a key role in their physiology. Human diets that are exclusively grain-based provide an excess of InsP6 that, through chelation of metal ions, may have a detrimental effect on human health. Ins(1,3,4,5,6)P5 2-kinase (InsP5 2-kinase or Ipk1) catalyses the synthesis of InsP6 from InsP5 and ATP, and is the only enzyme that transfers a phosphate group to the axial 2-OH of the myo-inositide. We present the first structure for an InsP5 2-kinase in complex with both substrates and products. This enzyme presents a singular structural region for inositide binding that encompasses almost half of the protein. The key residues in substrate binding are identified, with Asp368 being responsible for recognition of the axial 2-OH. This study sheds light on the unique molecular mechanism for the synthesis of the precursor of inositol pyrophosphates

Optimisation of the dibromomaleimide (DBM) platform for native antibody conjugation by accelerated post-conjugation hydrolysis

Disulfide bridging offers a convenient approach to generate site-selective antibody conjugates from native antibodies. To optimise the reagents available to achieve this strategy, we describe here the use of dibromomaleimides designed to undergo accelerated post-conjugation hydrolysis. Conjugation and hydrolysis, which serve to 'lock' the conjugates as robustly stable maleamic acids, is achieved in just over 1 h. This dramatic acceleration is also shown to infer significant improvements in homogeneity, as demonstrated by mass spectrometry analysis

Sulphur and carbon cycling in the subduction zone mélange

Subduction zones impose an important control on the geochemical cycling between the surficial and internal reservoirs of the Earth. Sulphur and carbon are transferred into Earth’s mantle by subduction of pelagic sediments and altered oceanic lithosphere. Release of oxidizing sulphate- and carbonate-bearing fluids modifies the redox state of the mantle and the chemical budget of subduction zones. Yet, the mechanisms of sulphur and carbon cycling within subduction zones are still unclear, in part because data are typically derived from arc volcanoes where fluid compositions are modified during transport through the mantle wedge. We determined the bulk rock elemental, and sulphur and carbon isotope compositions of exhumed ultramafic and metabasic rocks from Syros, Greece. Comparison of isotopic data with major and trace element compositions indicates seawater alteration and chemical exchange with sediment-derived fluids within the subduction zone channel. We show that small bodies of detached slab material are subject to metasomatic processes during exhumation, in contrast to large sequences of obducted ophiolitic sections that retain their seafloor alteration signatures. In particular, fluids circulating along the plate interface can cause sulphur mobilization during several stages of exhumation within high-pressure rocks. This takes place more pervasively in serpentinites compared to mafic rocks

Hand osteomyelitis in arterial calcification, diabetes mellitus and end-stage renal failure : a comparison of 210 cases over 12 years

We present 210 patients with hand osteomyelitis in 246 rays over 12 years, including detailed analysis of 29 patients in this cohort with digital artery calcification evident on plain X-ray. Overall 71 patients had diabetes mellitus and/or end-stage renal failure, including 28 of 29 patients with calcification. In the calcification group, 17 patients had ipsilateral arteriovenous fistulae, five had steal syndrome and 15 had digital ulceration or skin necrosis. Compared with 181 controls, patients with calcification had more affected bones, polymicrobial infections, surgical procedures, phalanges and digits amputated and had higher mortality at 1 year (12 of 29) and 5 years (20 of 29), as a result of comorbidities. Absence of calcification in 43 patients with diabetes and/or end-stage renal failure was associated with better outcomes on all the above parameters. Early amputation to maximize disease-free survival may be appropriate for patients with hand osteomyelitis and arterial calcification

TGFβ upregulates PAR-1 expression and signalling responses in A549 lung adenocarcinoma cells.

The major high-affinity thrombin receptor, proteinase activated receptor-1 (PAR-1) is expressed at low levels by the normal epithelium but is upregulated in many types of cancer, including lung cancer. The thrombin-PAR-1 signalling axis contributes to the activation of latent TGFβ in response to tissue injury via an αvβ6 integrin-mediated mechanism. TGFβ is a pleiotropic cytokine that acts as a tumour suppressor in normal and dysplastic cells but switches into a tumour promoter in advanced tumours. In this study we demonstrate that TGFβ is a positive regulator of PAR-1 expression in A549 lung adenocarcinoma cells, which in turn increases the sensitivity of these cells to thrombin signalling. We further demonstrate that this effect is Smad3-, ERK1/2- and Sp1-dependent. We also show that TGFβ-mediated PAR-1 upregulation is accompanied by increased expression of integrin αv and β6 subunits. Finally, TGFβ pre-stimulation promotes increased migratory potential of A549 to thrombin. These data have important implications for our understanding of the interplay between coagulation and TGFβ signalling responses in lung cancer.Medical Research Council UK (MRC) CASE studentship with Novartis awarded to RCC, MRC Centenary Award awarded to NS and RCC, and MRC Career Development Award G0800340 to CJS

How data visualisation using historical medical journals can contribute to current debates around antibiotic use and antimicrobial resistance in primary care

BackgroundThe early years of antibiotic use in primary care (c1950-1969) has received little attention. Medical journals provide a rich source for studying historic healthcare practitioners’ views and interests, with the potential to inform contemporary debate around issues of overuse and antimicrobial resistance. AimsPilot study to test the application of digital methods to interrogate historical medical journal data in relation to antibiotic use.Methods / ApproachMeta-data and scanned articles were extracted from the online British Journal of General Practice (BJGP) archive from inception (1953) to 1969. Searchable text was generated using an application called ABBYY optical character recognition, and Python used to generate data visualisations exploring (1) how BJGP changed during the period, (2) mentions of terms ‘antibiotic(s)’, ‘penicillin’, ‘resistance/resistant’ and mapping when and where they occurred.Results / EvaluationFrom 1953-1969, BJGP expanded in terms of number of annual issues (4 to 17) and annual pages (&lt;25 to &gt;1100). Heatmap visualisations were used to facilitate understanding of the frequency with which use of the term ‘antibiotic(s)’ occurred. By 1969 an article mentioning ‘antibiotic(s)’ was published monthly. Bigram searches found ‘treatment’ and ‘therapy’ to be the two most common terms that appeared with ‘antibiotic(s)’. The fourth and seventh most common terms were ‘resistant’ (first appearing in 1955) and ‘resistance’ (1962).ConclusionsThis pilot work shows that primary care publications increased considerably between 1953-1969. Articles on antibiotics featured frequently in relation to therapeutic intervention, and concerns around resistance occurred at an early stage. This approach provides new insights into how attitudes and behaviours around antibiotic use by primary care have evolved over time. It may also have the potential to inform study of the future use of antibiotics in primary care. <br/

A platform for efficient, thiol-stable conjugation to albumin's native single accessible cysteine

Herein we report the use of bromomaleimides for the construction of stable albumin conjugates via conjugation to its native, single accessible, cysteine followed by hydrolysis. Advantages over the classical maleimide approach are highlighted in terms of quantitative hydrolysis and absence of undesirable retro-Michael deconjugation

Serum inflammatory markers and amputations in hand osteomyelitis: a retrospective review of 146 cases

Background: The diagnosis of hand osteomyelitis requires correlation of clinical, radiological, and microbiological findings. The role of serum inflammatory markers in diagnosing and prognosticating hand osteomyelitis remains uncertain. We sought to determine the utility of inflammatory markers in the diagnosis and follow-up of hand osteomyelitis, and their ability to predict outcomes, particularly amputation. Methods: We retrospectively reviewed 146 patients diagnosed with hand osteomyelitis and with serum inflammatory marker levels measured after the onset of symptoms and within 14 days either side of diagnosis. Blood results at first presentation including white cell count (WCC), neutrophil-lymphocyte ratio (NLR), platelet-lymphocyte ratio (PLR), and C-reactive protein (CRP) were reviewed, and associations with amputations assessed. Follow-up markers taken at 15 to 60 days from diagnosis were analyzed where available. Results: Mean WCC and CRP at diagnosis were 9.2 (SD: 4.6) and 30.2 (SD: 42.4) respectively, compared with 8.2 (SD: 3.9) and 30.2 (SD: 42.4) at follow-up. At diagnosis, sensitivity of CRP was 74%, and WCC was 31%. Each marker had a low positive predictive value for amputation at diagnosis (<29%). A rise in CRP between diagnosis and follow-up was associated with an increased risk of amputation compared with a fall in CRP. The finding that WCC and CRP were both normal at diagnosis had a high negative predictive value against amputation (96%). Conclusion: C-reactive protein has a higher sensitivity than WCC, NLR, and PLR when used as a diagnostic adjunct in hand osteomyelitis. White cell count and CRP both within reference ranges at diagnosis was highly negatively predictive against amputation

Functionalized Cyclopentenones with Low Electrophilic Character as Anticancer Agents

In this study were synthesized non-Michael acceptor cyclopentenones (CP) from biomass derivative furfural as anticancer agents. Cyclic enones, both from natural sources and synthetic analogues, have been described as cytotoxic agents. Most of these agents were unsuccessful in becoming valuable therapeutic agents due to toxicity problems derived from unselective critical biomacromolecule alkylation. This may be caused by Michael addition to the enone system. Ab initio studies revealed that 2,4-substituted CPs are less prone to Michael additions, and as such were tested three families of those derivatives. We prepare the new CPs from furfural through a tandem furan ring opening/nazarov electrocyclization and further functionalization. Experimentally the 2,4-substituted CPs exhibited no reactivity towards sulphur nucleophiles, while maintaining cytotoxicity against HT-29, MCF-7, NCI-H460, HCT-116 and MDA-MB 231 cells lines. Moreover, the selected CP are non-toxic against healthy HEK 293T cell lines and present proper calculated drug-like properties