Heat-induced changes in the finite strain viscoelastic behavioir of a collaagenous tissue

Supra-physiological temperatures are increasingly being used to treat many different soft need for injuries. To identify improved clinical treatments, however, there is a need for better information on the effect of the mechanics on the thermal damage process as well as the effect of the incurred damage on the subsequent mechanical properties. In this paper we report the first biaxial data on the stress relaxation behavior of a collagenous tissue before and after thermal damage. Based on a two-dimensional finite strain viscoelastic model, which incorporates an exponential elastic response, it is shown that the thermal damage can significantly decrease the characteristic time for stress relaxation and the stress residual.     

Survey of extreme solvent tolerance in gram-positive cocci: membrane fatty acid changes in Staphylococcus haemolyticus grown in toluene

We exploited the unique ecological niche of oil fly larval guts to isolate a strain of Staphylococcus haemolyticus which may be the most solvent-tolerant gram-positive bacterium yet described. This organism is able to tolerate 100% toluene, benzene, and p-xylene on plate overlays and saturating levels of these solvents in monophasic liquid cultures. A comparison of membrane fatty acids by gas chromatography after growth in liquid media with and without toluene showed that in cells continuously exposed to solvent the proportion of anteiso fatty acids increased from 25.8 to 33.7% while the proportion of 20:0 straight-chain fatty acids decreased from 19.3 to 10.1%. No changes in the membrane phospholipid composition were noted. Thus, S. haemolyticus alters its membrane fluidity via fatty acid composition to become more fluid when it is exposed to solvent. This response is opposite that commonly found in gram-negative bacteria, which change their fatty acids so that the cytoplasmic membrane is less fluid. Extreme solvent tolerance in S. haemolyticus is not accompanied by abnormal resistance to anionic or cationic detergents. Finally, six strains of Staphylococcus aureus and five strains of Staphylococcus epidermidis, which were not obtained by solvent selection, also exhibited exceptional solvent tolerance.     

Model of Vibrio cholerae toxin coregulated pilin capable of filament formation

A complete three-dimensional model (RCSB001169; PDB code 1qqz ) for the Vibrio cholerae toxin coregulated pilus protein (TcpA), including residues 1-197, is presented. We have used the crystal structure of the Neisseria gonorrhoeae pilin (PilE), available biochemical data about TcpA, variations in the primary sequences of TcpA among various Vibrio cholerae strains and secondary structure prediction, hydrophilicity, surface probability and antigenicity plots for TcpA to build our model. In our TcpA model, the first 137 residues possess a structure similar to the PilE, but the remainder is different. Though the ladle shape is still preserved, TcpA possesses a larger ladle head or globular domain compared to PilE. Using this model, it has been possible to identify two kinds of conserved residues: (i) those forming the core of the TcpA monomer and (ii) those involved in the monomer-monomer interactions leading to fibre formation. Residues on the fibre exterior, important in the mediation of bacterium (pilus)-bacterium (pilus) and bacterium (pilus)-host interactions, show more variability in comparison to those of (i) and (ii).     

Substitution of Met-69 by Ala or Gly in TEM-1 beta-lactamase confer an increased susceptibility to clavulanic acid and other inhibitors

In some inhibitor-resistant TEM-derived beta-lactamases, Met-69 is substituted by Leu, Ile or Val. Residue 69 is located in a region of strong structural constraints, at the beginning of H2 alpha-helix, and in the vicinity of B3 and B4 beta-strands. Analysis of the three-dimensional structure of TEM-1 beta-lactamase suggests that alteration of the substrate-binding site can be produced by changes of the size of residue 69 side chain. Met-69 was substituted by alanine or glycine in TEM-Bs beta-lactamase (a TEM-1-related enzyme) using site-directed mutagenesis. The minimum inhibitory concentrations of the mutants compared with the wild-type revealed an increased susceptibility to beta-lactamase inhibitor-beta-lactam combinations and to first-generation cephalosporins. Comparing the Met69Ala and Met69Gly beta-lactamases with TEM-Bs, K(m) constants of the mutants showed an increased affinity for most beta-lactams but the kcat for most substrates did not change substantially. Mutants also demonstrated lower IC50 for the three inhibitors (clavulanic acid, tazobactam and sulbactam). The two substitutions of the residue 69 by alanine and glycine had a noticeable effect on K(m) values of TEM-Bs beta-lactamase, and on affinity for beta-lactamase inhibitors.     

Sodium dodecyl sulfate hypersensitivity of clpP and clpB mutants of Escherichia coli

We studied the hypersensitivity of clpP and clpB mutants of Escherichia coli to sodium dodecyl sulfate (SDS). Both wild-type E. coli MC4100 and lon mutants grew in the presence of 10% SDS, whereas isogenic clpP and clpB single mutants could not grow above 0.5% SDS and clpA and clpX single mutants could not grow above 5.0% SDS. For wild-type E. coli, cellular ClpP levels as determined by Western immunoblot analysis increased ca. sixfold as the levels of added SDS increased from 0 to 2%. Capsular colanic acid, measured as uronic acid, increased ca. sixfold as the levels of added SDS increased from 2 to 10%. Based on these findings, 3 of the 19 previously identified SDS shock proteins (M. Adamowicz, P. M. Kelley, and K. W. Nickerson, J. Bacteriol. 173:229-233, 1991) are tentatively identified as ClpP, ClpX, and ClpB.     

Biological investigation and structure-activity relationship studies on azadirone from Azadirachta indica A. Juss

Azadirone 1, a limonoidal constituent of Azadirachta indica is found to possess potent cytotoxic activity against a panel of human cancer cell lines in our in vitro studies. In vitro screening of a number of semi-synthetic analogues of 1 revealed that the alpha,beta-unsaturated enone moiety or its equivalent conjugated system in A-ring, C-7 acetyloxy/chloroacetyloxy or keto group in B-ring and the furan moiety are responsible for the activity of 1 and its analogues. Compound 1 and two of the semi-synthetic analogues 10 and 13 were found to possess good in vivo antitumor activity in modified hollow fiber animal models.     

Novel 6,7-diphenyl-2,3,8,8a-tetrahydro-1H-indolizin-5-one analogues as cytotoxic agents

A series of 6,7-diphenyl-2,3,8,8a-tetrahydro-1H-indolizin-5-one analogues were synthesized and evaluated for cytotoxic activity against eight human cancer cell lines. Compounds 18, 21, 28, 29, 30 and 31 showed cytotoxic activity with GI(50) values in the range of 2.1-8.1 microM concentration. Among these, compounds 21 and 28 exhibited good pharmacokinetic properties. These compounds were further evaluated for their in vivo efficacy in modified hollow fibre assay (HFA).     

Rab5-mediated endosome-endosome fusion regulates hemoglobin endocytosis in Leishmania donovani

To understand the trafficking of endocytosed hemoglobin (Hb) in Leishmania, we investigated the characteristics of in vitro fusion between endosomes containing biotinylated Hb (BHb) and avidin-horseradish peroxidase (AHRP). We showed that early endosome fusion in Leishmania is temperature and cytosol dependent and is inhibited by ATP depletion, ATPgammaS, GTPgammaS and N-ethylmaleimide treatment. The Rab5 homolog from Leishmania donovani, LdRab5, was cloned and expressed. Our results showed that homotypic fusion between the early endosomes in Leishmania is Rab5 dependent. Early endosomes containing BHb fused efficiently with late endosomes in a process regulated by Rab7, whereas no fusion between early and late endosomes was detected using fluid phase markers. Pre-treatment of early endosomes containing BHb with monoclonal antibody specific for the C-terminus of the Hb receptor (HbR) or the addition of the C-terminal cytoplasmic fragment of the HbR specifically inhibited the fusion with late endosomes, suggesting that signal(s) mediated through the HbR cytoplasmic tail promotes the fusion of early endosomes containing Hb with late endosomes.