Experimental Validation of the Predicted Binding Site of Escherichia coli K1 Outer Membrane Protein A to Human Brain Microvascular Endothelial Cells: Identification of Critical Mutations That Prevent E. coli Meningitis

Escherichia coli K1, the most common cause of meningitis in neonates, has been shown to interact with GlcNAc1–4GlcNAc epitopes of Ecgp96 on human brain microvascular endothelial cells (HBMECs) via OmpA (outer membrane protein A). However, the precise domains of extracellular loops of OmpA interacting with the chitobiose epitopes have not been elucidated. We report the loop-barrel model of these OmpA interactions with the carbohydrate moieties of Ecgp96 predicted from molecular modeling. To test this model experimentally, we generated E. coli K1 strains expressing OmpA with mutations of residues predicted to be critical for interaction with the HBMEC and tested E. coli invasion efficiency. For these same mutations, we predicted the interaction free energies (including explicit calculation of the entropy) from molecular dynamics (MD), finding excellent correlation (R^2 = 90%) with experimental invasion efficiency. Particularly important is that mutating specific residues in loops 1, 2, and 4 to alanines resulted in significant inhibition of E. coli K1 invasion in HBMECs, which is consistent with the complete lack of binding found in the MD simulations for these two cases. These studies suggest that inhibition of the interactions of these residues of Loop 1, 2, and 4 with Ecgp96 could provide a therapeutic strategy to prevent neonatal meningitis due to E. coli K1

Regional nitrogen cycle: an Indian perspective

During the past century through food and energy production, human activities have altered the world's nitrogen cycle by accelerating the rate of reactive nitrogen creation. India has made impressive strides in the agricultural front, in which N fertilizer plays a major role. There has been a marked change in the supply and use of land, water, fertilizers, seeds and livestock, but the N use efficiency remained at a low level. Exploring the nature of these changes and quantification of the impacts on the N cycle has become essential. Hence we have presented data on various N pools and fluxes based on a conceptual N model. In India, efforts should focus on understanding the fate and consequences of the applied N and to increase the efficiency of N use

Nitrate content in wheat leaf blades

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,3-Sigmatropic fluorine migration to boron in McLafferty type of rearrangements: Observation of tetrafluorobenzyne radical cation and trifluorobenzyne cation by CID-mass spectrometry

a b s t r a c t The electron-impact mass spectra of the tris(pentafluorophenyl)boroxine (1) and triphenylboroxine (2) were analyzed to probe the McLafferty type of rearrangements involving 1,3-fluorine or 1,3-hydrogen migrations to boron from the adjacent aromatic rings. 1,3-Sigmatropic fluorine migration leading to the McLafferty rearrangement of 1 results in the formation of the tetrafluorobenzyne radical cation (1c), whereas the similar rearrangement involving 1,3-hydrogen rearrangement is not observed for the nonfluorinated analog 2. The DFT calculations show that the activation barrier for the 1,3-fluorine migration in 1 is significantly lower than that for the 1,3-hydrogen migration in 2 (DDG z w33 kcal/ mol), which is in accordance with the observed 1,3-fluorine migration for the fluorinated boroxine and the lack of such rearrangement for the nonfluorinated boroxine. The 1:1 stoichiometry of the fluoride anion with 1 has also been demonstrated by high resolution electrospray ionization time-of-flight mass spectrometry

Novel purine-based fluoroaryl-1,2,3-triazoles as neuroprotecting agents: Synthesis, neuronal cell culture investigations, and CDK5 docking studies

A series of novel purine-based fluoroaryl triazoles were synthesized using the Cu(I) catalyzed 1,3-dipolar cycloaddition reactions (click reactions), and assayed for their neuroprotective effects using fluorescence electron microscopy. Among these triazoles, o-fluorophenylmetyl-triazole, 7, has comparable neuroprotective effect as that of Flavopiridol (1) and Roscovitine (2), the state of the art CDK inhibitors, against the Aβ induced neurotoxicity. These results are substantiated using computer docking methods (DarwinDock/GenDock), which predict that Roscovitine and the triazole 7 bind to the ATP-binding site of CDK5/p25 with comparable binding energies, whereas the corresponding pentafluorophenylmethyl-triazole, 9, has dramatically reduced binding energy (in accordance with its lack of neuroprotection). These combined experimental and theoretical studies support the involvement of CDK5/p25 in the neuronal cell cycle re-entry

Evidence-based national vaccine policy

India has over a century old tradition of development and production of vaccines. The Government rightly adopted self-sufficiency in vaccine production and self-reliance in vaccine technology as its policy objectives in 1986. However, in the absence of a full-fledged vaccine policy, there have been concerns related to demand and supply, manufacture vs. import, role of public and private sectors, choice of vaccines, new and combination vaccines, universal vs. selective vaccination, routine immunization vs. special drives, cost-benefit aspects, regulatory issues, logistics etc. The need for a comprehensive and evidence based vaccine policy that enables informed decisions on all these aspects from the public health point of view brought together doctors, scientists, policy analysts, lawyers and civil society representatives to formulate this policy paper for the consideration of the Government. This paper evolved out of the first ever ICMR-NISTADS national brainstorming workshop on vaccine policy held during 4-5 June, 2009 in New Delhi, and subsequent discussions over email for several weeks, before being adopted unanimously in the present form