Instability of aquaglyceroporin (Aqp) 2 contributes to drug resistance in trypanosoma brucei

Defining mode of action is vital for both developing new drugs and predicting potential resistance mechanisms. Sensitivity of African trypanosomes to pentamidine and melarsoprol is predominantly mediated by aquaglyceroporin 2 (TbAQP2), a channel associated with water/glycerol transport. TbAQP2 is expressed at the flagellar pocket membrane and chimerisation with TbAQP3 renders parasites resistant to both drugs. Two models for how TbAQP2 mediates pentamidine sensitivity have emerged; that TbAQP2 mediates pentamidine translocation across the plasma membrane or via binding to TbAQP2, with subsequent endocytosis and presumably transport across the endosomal/lysosomal membrane, but as trafficking and regulation of TbAQPs is uncharacterised this remains unresolved. We demonstrate that TbAQP2 is organised as a high order complex, is ubiquitylated and is transported to the lysosome. Unexpectedly, mutation of potential ubiquitin conjugation sites, i.e. cytoplasmic-oriented lysine residues, reduced folding and tetramerization efficiency and triggered ER retention. Moreover, TbAQP2/TbAQP3 chimerisation, as observed in pentamidine-resistant parasites, also leads to impaired oligomerisation, mislocalisation and increased turnover. These data suggest that TbAQP2 stability is highly sensitive to mutation and that instability contributes towards the emergence of drug resistance

Contaminants of post-leaching copper solutions and their behavior during extraction with industrial extractants

Several copper extractants have been examined in laboratory glassware with a special attention paid to their behavior towards cations others than copper(II). In the studies 25 vol. percent of hydrocarbon (Exxsol D80 AZ) solutions of six industrial copper extractants have been used. They were mainly rea-gents of hydrooxime type (LIX 860N-IC, LIX 984, LIX 984N, LIX 84-I, Acorga M5640) and diketone type LIX 54-100. Individual isotherms of independent extraction of copper(II) and selected cations (Fe3+, Fe2+, Co2+, Ni2+, Zn2+, Mn2+, Cd2+, Mg2+) versus equilibrium pH have been compared. Investigations have been conducted with synthetic 0.1 mol/L solutions of their sulfates. The values of pH50% − parameter defined as an equilibrium pH at the moment of half-and-half extraction of the investigated cationic spe-cies have been presented. Also ΔpH50%, that is differences between values of pH50% for specific cation Me(II) and copper(II): ΔpH50% = pH50%(Me) − pH50%(Cu), have been given. In addition to that 25% LIX 984 has been used in counter-current pilot trials for copper(II) extraction from naturally contaminated solutions produced by bioleaching of industrial sulfide copper concentrate to observe behavior of investi-gated contaminants such as correlations between their real co-extraction with copper(II) and the position of their extraction on the pH scale. Copper electrolyte/strip solution, working alternately within close loop of coupled stripping - electrowinning system, has been analyzed during consecutive cycles to observe building up of the contaminants concentrations in the course of test

Recovery of major components of spent zinc electrolyte with di(2-ethylhexyl)phosphoric acid (DEHPA) used as a zinc(II) extractant

Industrial acidic zinc electrolyte has been treated with calcium compounds and/or concentrated ammonia solution and resulting crystalline phases as well as equilibrium mother liquors were analyzed in detail for dependence on preliminary dilution of electrolyte with water. Neutralized zinc electrolyte was an object of investigation in zinc(II) extraction with di(2-ethylhexyl)phosphoric acid (DEHPA) as an extractant. Dependencies of zinc(II) and contaminants extraction on equilibrium pH, isotherms of zinc(II) extraction and stripping have been studied. During simulated laboratory counter-current trial of zinc extraction a new zinc electrolyte (stripped solution) has been produced. Raffinates left after the process were purified and qualified as mineral magnesium-ammonium fertilizer solutions

Direct measurements of reaction time for extractive processes

Extraction times for selected metal cations (Mg2+, Al3+, Zn2+, Cd2+, Cu2+, Ni2+, Co2+, Mn2+, Fe2+ and Fe3+) have been measured using cation-exchanging extractants - Versatic 10 (neodecanoic acid), DEHPA (bis(2-ethylhexyl)phosphoric acid) and Cyanex 272 (bis(2,4,4-trimetylpentyl)phosphinic acid). Data was collected directly by measuring and recording pH of the two phase reaction mixture with maintaining continuity of the aqueous phase. Similarly, the stripping rate of selected cations (Mg2+, Al3+, Zn2+, Cu2+, Ni2+, Co2+) in a sulphuric acid solution was measured. Viscosities of organic phases, saturated with cations, were measured before their stripping. Additionally, the extraction dependence of Mg2+, Al3+, Fe2+ and Fe3+ on pH was measured using Versatic 10 at various concentrations (7.5, 15 and 30 %)

Neodecanoic acid as extractant of selected non-ferrous metals

The paper presents results of studies on extracting properties of neodecanoic acid (Versatic 10) solutions. Neodecanoic acid was diluted in Exxsol D80 AZ to three concentrations: 7.5, 15, and 30 % (v/v). Tests were conducted using sulphate(VI) solutions of six ions: Zn(II), Ni(II), Cu(II), Co(II), Cd(II), Mn(II) and a nitrate(V) solution of Pb(II). All the aqueous solutions were prepared as 0.1 M concentration. For all combinations of the used solutions (organic and aqueous), direct methods of measuring and recording the pH of the biphasic mixture have been applied. The degrees of ions extraction, depending on equilibrium pH were also presented in the paper. Extraction and stripping isotherms of selected ions and organic phase were plotted for 7.5 and 30 % (v/v) of extractant solutions and Zn(II), Ni(II) and Cu(II) ions

Strengthening the response to disasters and trauma

Chapter 10 opens with the Gulf Research Program’s Lauren Alexander Augustine presenting four pilot programs from around the country that illustrate the cornerstones of community resilience. Next, Benjamin Springgate examines resilience as a reflection of individual communities and describes a research network that promotes research on resilience-strengthening practices. Lourdes J. Rodríguez and Sheila B. Savannah then explain how neighborhoods that are striving for resilience should be involved like patients in the process of healing. Finally, three contributors offer unique perspectives on resilient communities: Traci L. Birch looks at why resilience efforts should also focus on inland communities that are no longer immune to the impacts of climate change and extreme weather. And Joie B. Acosta and Elka Gotfryd encourage us to think broadly about resilience as they explore opportunities to build responsive social infrastructure through engaged citizen participation

Mechanism of the Association between Na+Binding and Conformations at the Intracellular Gate in Neurotransmitter:Sodium Symporters

Neurotransmitter:sodium symporters (NSSs) terminate neurotransmission by Na(+)-dependent reuptake of released neurotransmitters. Previous studies suggested that Na(+)-binding reconfigures dynamically coupled structural elements in an allosteric interaction network (AIN) responsible for function-related conformational changes, but the intramolecular pathway of this mechanism has remained uncharted. We describe a new approach for the modeling and analysis of intramolecular dynamics in the bacterial NSS homolog LeuT. From microsecond-scale molecular dynamics simulations and cognate experimental verifications in both LeuT and human dopamine transporter (hDAT), we apply the novel method to identify the composition and the dynamic properties of their conserved AIN. In LeuT, two different perturbations disrupting Na(+) binding and transport (i.e. replacing Na(+) with Li(+) or the Y268A mutation at the intracellular gate) affect the AIN in strikingly similar ways. In contrast, other mutations that affect the intracellular gate (i.e. R5A and D369A) do not significantly impair Na(+) cooperativity and transport. Our analysis shows these perturbations to have much lesser effects on the AIN, underscoring the sensitivity of this novel method to the mechanistic nature of the perturbation. Notably, this set of observations holds as well for hDAT, where the aligned Y335A, R60A, and D436A mutations also produce different impacts on Na(+) dependence. Thus, the detailed AIN generated from our method is shown to connect Na(+) binding with global conformational changes that are critical for the transport mechanism. That the AIN between the Na(+) binding sites and the intracellular gate in bacterial LeuT resembles that in eukaryotic hDAT highlights the conservation of allosteric pathways underlying NSS function