Disease-causing point-mutations in metal-binding domains of Wilson disease protein decrease stability and increase structural dynamics

After cellular uptake, Copper (Cu) ions are transferred from the chaperone Atox1 to the Wilson disease protein (ATP7B) for incorporation into Cu-dependent enzymes in the secretory pathway. Human ATP7B is a large multi-domain membrane-spanning protein which, in contrast to homologues in other organisms, has six similar cytoplasmic metal-binding domains (MBDs). The reason for multiple MBDs is proposed to be indirect modulation of enzymatic activity and it is thus intriguing that point mutations in MBDs can promote Wilson disease. We here investigated, in vitro and in silico, the biophysical consequences of clinically-observed Wilson disease mutations, G85V in MBD1 and G591D in MBD6, incorporated in domain 4. Because G85 and G591 correspond to a conserved Gly found in all MBDs, we introduced the mutations in the well-characterized MBD4. We found the mutations to dramatically reduce the MBD4 thermal stability, shifting the midpoint temperature of unfolding by more than 20 °C. In contrast to wild type MBD4 and MBD4D, MBD4V adopted a misfolded structure with a large β-sheet content at high temperatures. Molecular dynamic simulations demonstrated that the mutations increased backbone fluctuations that extended throughout the domain. Our findings imply that reduced stability and enhanced dynamics of MBD1 or MBD6 is the origin of ATP7B dysfunction in Wilson disease patients with the G85V or G591D mutation.     

Interactions between allelopathic properties and growth kynetics in four freshwater phytoplankton species studied by model simulations

We chose four species of freshwater phytoplankton: the chlorophyceans Ankistrodesmus falcatus, Chlamydomonas reinhardtii and Selenastrum capricornutum, and the cyanobacteria Oscillatoria sp. in order to study their competitive abilities for nitrate and their allelopathic properties. We parameterized models of nitrate uptake and growth with laboratory experiments. According to them, the species were ranked (from the best to the worst competitors): S. capricornutum, C. reinhardtii, A. falcatus and Oscillatoria sp. C. reinhardtii and Oscillatoria sp. were previously reported as allelopathic. In the present work, Oscillatoria sp. was allelopathic only against A. falcatus. However, none of our species was sensitive to C. reinhardtii. Additionally, we found an unknown allelopathic effect of A. falcatus against Oscillatoria sp. Our findings point out the high specificity of allelopathic interactions. With these data, we constructed a model of interspecific competition for nitrate, including allelopathic interactions. By performing model simulations, we studied how three factors influence the outcome of competition: relative abundance of competing species, resistance to allelopathy, and nitrate concentration. Our simulations showed that the initial ratio of species abundances will significantly determine the outcome of competition. If the worst competitor was the allelopathic species, the more it needs to outnumber the competing species, unless it is very sensitive to allelopathy (not defended). Nitrate has an important influence, showing a non-intuitive outcome of competition experiments at low nitrate concentrations, where the worst competitor (allelopathic species) wins competition in the majority of cases, whereas at intermediate concentrations, the better competitor dominates except for unfavorable ratios of abundances. With the increased amounts of nitrate, conditions again favor the worst competitor (the stronger allelopathic species). Despite the potential for two species coexistence showed by previous theoretical analysis of systems was similar to ours, our simulations did not detect this outcome. We hypothesized that this is due to the strong allelopathic effect of Oscillatoria sp.     

Are there risk factors for acute renal failure in adult patients using deoxycholate amphotericin B?

BackgroundDeoxycholate amphotericin B (DAB) is a nephrotoxic drug and the incidence of acute kidney injury (AKI) is high.AimsThe aim of this study was to describe the incidence of AKI in patients under DAB therapy and determine risk factor to predict the AKI.MethodsThe data of this retrospective study included previously hospitalized patients treated with intravenous DAB for at least five days. Clinical and laboratorial data were evaluated and AKI was classified in stages using Acute Kidney Injury Network criteria. Univariated test followed by a multivariable analysis was performed to determine risk factor and Kaplan–Meier survival estimates were calculated to evaluate the role of AKI in the outcome.ResultsOne hundred six patients were included in the final analysis. AKI occurred in 51.9% and dialysis was necessary in 4.7%. The occurrence of AKI was not associated with any risk factor. The mortality of the patients was neither associated with AKI nor with dialysis. Other nephrotoxic drugs were not risk factors for AKI.ConclusionsThe incidence of AKI in patients using DAB is high and we cannot predict the chance of AKI using clinical or laboratorial data.     

A C solid state nuclear magnetic resonance spectroscopic study of cork cell wall structure: the effect of suberin removal

Solid state ¹³C NMR measurements of cork, before and after suberin removal, showed that aliphatic suberin is spatially separated from carbohydrate and lignin and experiences higher motional freedom. Two types of chain methylenes, differing in chemical shift and in dynamic properties, were identified in aliphatic suberin. Experimental evidence indicated that the more motionally hindered methylenes are those situated nearer the linkages of aliphatic suberin to the cell wall. These linkages were shown to involve –CH₂O– groups, probably engaged in ester linkages to phenylpropane units and carbohydrate C6 carbons. Spectral intensity changes indicated that, during the first steps of alkaline desuberization, these linkages are broken and the shorter aliphatic suberin chains removed. Longer chains require hydrolysis of the ester linkages within the chains and are removed upon stronger alkaline treatment. T₁(C), T_1ρ(H) and T_1ρ(C) relaxation times have shown that the removal of suberin from cork leads to a motionally restricted and more compact environment, on the megahertz and mid-kilohertz timescales. The properties of cork suberin showed that suberin organization in cork is distinct from that in potato tissue.     

Metabolic engineering strategies for butanol production in Escherichia coli

The global market of butanol is increasing due to its growing applications as solvent, flavoring agent, and chemical precursor of several other compounds. Recently, the superior properties of n-butanol as a biofuel over ethanol have stimulated even more interest. (Bio)butanol is natively produced together with ethanol and acetone by Clostridium species through acetone-butanol-ethanol fermentation, at noncompetitive, low titers compared to petrochemical production. Different butanol production pathways have been expressed in Escherichia coli, a more accessible host compared to Clostridium species, to improve butanol titers and rates. The bioproduction of butanol is here reviewed from a historical and theoretical perspective. All tested rational metabolic engineering strategies in E. coli to increase butanol titers are reviewed: manipulation of central carbon metabolism, elimination of competing pathways, cofactor balancing, development of new pathways, expression of homologous enzymes, consumption of different substrates, and molecular biology strategies. The progress in the field of metabolic modeling and pathway generation algorithms and their potential application to butanol production are also summarized here. The main goals are to gather all the strategies, evaluate the respective progress obtained, identify, and exploit the outstanding challenges.