Investigating the anti-inflammatory and analgesic properties of leaves ethanolic extracts of Cedrus libani and Pinus brutia.

This investigation aimed to assess the anti-inflammatory and analgesic effects of Cedrus libani and Pinus brutia leaves. The anti-inflammatory property was evaluated by Human Red Blood Cells (HRBC) membrane stabilization assay and Albumin denaturation assay using Sodium diclofenac as a positive control. To evaluate the analgesic property, formalin and tail flick tests were carried out using ethanolic extracts at a dose of 30 mg/kg and gel containing 2% (w/v) of ethanolic extract of each plant. Diclofenac sodium, diclofenac gel 1% and lidocaine gel 2 % were used as positive controls. Results: The effect of inhibiting hemolysis was observed at concentrations (2.5-12.5) µg/ml for P. brutia, and (2.5-25) µg/ml for C. libani. Moreover, albumin denaturation test showed protection effect for both plant extracts with IC50 of 47.74 and 81.50 µg/ml for C. libani and P. brutia extract, consecutively. In Formalin test, both extracts could significantly reduce paw licking time, and in tail flick test, each plant extract gel showed greater efficacy than diclofenac gel by calculating the maximum possible effect (MPE %) for both extracts and Diclofenac. Conclusion: We concluded that both extracts showed in vitro anti-inflammatory activity at different concentrations when compared to standard drug of diclofenac as well as analgesic activity in formalin and tail flick tests.

TraG encoded by the pIP501 type IV secretion system is a two-domain peptidoglycan-degrading enzyme essential for conjugative transfer.

pIP501 is a conjugative broad-host-range plasmid frequently present in nosocomial Enterococcus faecalis and Enterococcus faecium isolates. We focus here on the functional analysis of the type IV secretion gene traG, which was found to be essential for pIP501 conjugative transfer between Gram-positive bacteria. The TraG protein, which localizes to the cell envelope of E. faecalis harboring pIP501, was expressed and purified without its N-terminal transmembrane helix (TraGΔTMH) and shown to possess peptidoglycan-degrading activity. TraGΔTMH was inhibited by specific lytic transglycosylase inhibitors hexa-N-acetylchitohexaose and bulgecin A. Analysis of the TraG sequence suggested the presence of two domains which both could contribute to the observed cell wall-degrading activity: an N-terminal soluble lytic transglycosylase domain (SLT) and a C-terminal cysteine-, histidine-dependent amidohydrolases/peptidases (CHAP) domain. The protein domains were expressed separately, and both degraded peptidoglycan. A change of the conserved glutamate residue in the putative catalytic center of the SLT domain (E87) to glycine resulted in almost complete inactivity, which is consistent with this part of TraG being a predicted lytic transglycosylase. Based on our findings, we propose that TraG locally opens the peptidoglycan to facilitate insertion of the Gram-positive bacterial type IV secretion machinery into the cell envelope.

Crystallization and first data collection of the putative transfer protein TraN from the Gram-positive conjugative plasmid pIP501.

Conjugative plasmid transfer is the most important route for the spread of resistance and virulence genes among bacteria. Consequently, bacteria carrying conjugative plasmids are a substantial threat to human health, especially hospitalized patients. Whilst detailed information about the process has been obtained for Gram-negative type-4 secretion systems, little is known about the corresponding mechanisms in Gram-positive (G+) bacteria. The successful purification and crystallization of the putative transfer protein TraN from the G+ conjugative model plasmid pIP501 of Enterococcus faecalis are presented. Native crystals diffracted to 1.8 Šresolution on a synchrotron beamline. The crystals belonged to space group P2(1), with unit-cell parameters a=32.88, b=54.94, c=57.71 Å, ß=91.89° and two molecules per asymmetric unit.