Oligomers of Heat-Shock Proteins: Structures That Don’t Imply Function

Most proteins must remain soluble in the cytosol in order to perform their biological functions. To protect against undesired protein aggregation, living cells maintain a population of molecular chaperones that ensure the solubility of the proteome. Here we report simulations of a lattice model of interacting proteins to understand how low concentrations of passive molecular chaperones, such as small heat-shock proteins, suppress thermodynamic instabilities in protein solutions. Given fixed concentrations of chaperones and client proteins, the solubility of the proteome can be increased by tuning the chaperone–client binding strength. Surprisingly, we find that the binding strength that optimizes solubility while preventing irreversible chaperone binding also promotes the formation of weakly bound chaperone oligomers, although the presence of these oligomers does not significantly affect the thermodynamic stability of the solution. Such oligomers are commonly observed in experiments on small heat-shock proteins, but their connection to the biological function of these chaperones has remained unclear. Our simulations suggest that this clustering may not have any essential biological function, but rather emerges as a natural side-effect of optimizing the thermodynamic stability of the proteome.     

Is land use impact assessment in LCA applicable for forest biomass value chains? Findings from comparison of use of Scandinavian wood,agro-biomass and peat for energy

Purpose

A framework for the inclusion of land use impact assessment and a set of land use impact indicators has been recently proposed for life cycle assessment (LCA) and no case studies are available for forest biomass. The proposed methodology is tested for Scandinavian managed forestry; a comparative case study is made for energy from wood, agro-biomass and peat; and sensitivity to forest management options is analysed.

Methods

The functional unit of this comparative case study is 1 GJ of energy in solid fuels. The land use impact assessment framework of the United Nations Environment Programme and the Society of Environmental Toxicology and Chemistry (UNEP-SETAC) is followed and its application for wood biomass is critically analysed. Applied midpoint indicators include ecological footprint and human appropriation of net primary production, global warming potential indicator for biomass (GWP_bio-100) and impact indicators proposed by UNEP-SETAC on ecosystem services and biodiversity. Options for forest biomass land inventory modelling are discussed. The system boundary covers only the biomass acquisition phase. Management scenarios are formulated for forest and barley biomass, and a sensitivity analysis focuses on impacts of land transformations for agro-biomass.

Results and discussion

Meaningful differences were found in between solid biofuels from distinct land use classes. The impact indicator results were sensitive to land occupation and transformation and differed significantly from inventory results. Current impact assessment method is not sensitive to land management scenarios because the published characterisation factors are still too coarse and indicate differences only between land use types. All indicators on ecosystem services and biodiversity were sensitive to the assumptions related with land transformation. The land occupation (m²a) approach in inventory was found challenging for Scandinavian wood, due to long rotation periods and variable intensities of harvests. Some suggestions of UNEP-SETAC were challenged for the sake of practicality and relevance for decision support.

Conclusions

Land use impact assessment framework for LCA and life cycle impact assessment (LCIA) indicators could be applied in a comparison of solid bioenergy sources. Although forest bioenergy has higher land occupation than agro-bioenergy, LCIA indicator results are of similar magnitude or even lower for forest bioenergy. Previous literature indicates that environmental impacts of land use are significant, but it remains questionable if these are captured with satisfactory reliability with the applied LCA methodology, especially for forest biomass. Short and long time perspectives of land use impacts should be studied in LCA with characterisation factors for all relevant timeframes, not only 500 years, with a forward-looking perspective. Characterisation factors need to be modelled further for different (forest) land management intensities and for peat excavation.     

A recent local sweep at the PHYA locus in the Northern European Spiterstulen population of Arabidopsis lyrata

Northern and central European Arabidopsis lyrata ssp. petraea populations are locally adapted to prevailing climatic conditions through differences in timing of life history events. The timing of flowering and, in perennials, the timing of growth cessation influence fitness. Phytochrome A may have an important role in regulating these life history traits as it perceives changes in daylength. We asked whether PHYA has contributed to local adaptation to the northern conditions in A. l. petraea. To search for signals of directional selection at the PHYA locus, we resequenced PHYA and 9 short fragments around PHYA from a 57‐kb region from a German (Plech) and a Norwegian (Spiterstulen) population and compared patterns of differentiation and diversity to a set of 19 reference loci around the genome. First, we found that the populations were highly differentiated: there were three nonsynonymous fixed differences at the PHYA locus, which was in stark contrast with the total four fixed differences in the 19 reference loci. Compatible with a sweep hypothesis, variation was almost completely removed from the 9.4‐kb region around PHYA in the northern Spiterstulen population. The overall level of linkage disequilibrium (LD) was higher in Spiterstulen, but there was no LD across the PHYA locus in the population, which is also a known consequence of a selective sweep. The sweep has likely occurred after the last glacial maximum, which suggests that it has contributed to adaptation to the northern conditions.     

Chemical models for ribozyme action

Mechanistic studies of the action of catalytic ribonucleic acids, ribozymes, are highly challenging, because even a slight structural change can dramatically affect the chain folding. This, in turn, alters the binding properties of the catalytic core, making identification of the real origin of the observed influence on rate difficult. Unambiguous structure-reactivity correlations based on studies with structurally simplified chemical models may help to distinguish between alternative mechanistic interpretations. The results of such model studies are reviewed. The topics include intramolecular cleavage of RNA phosphodiester bonds by solvent-derived species, general acids/bases and metal ions, effect of molecular environment on their hydrolytic stability and trinucleoside monophosphates as models for large ribozymes.     

Functional genes reveal the intrinsic PAH biodegradation potential in creosote-contaminated groundwater following in situ biostimulation

A small-scale functional gene array containing 15 functional gene probes targeting aliphatic and aromatic hydrocarbon biodegradation pathways was used to investigate the effect of a pilot-scale air sparging and nutrient infiltration treatment on hydrocarbon biodegradation in creosote-contaminated groundwater. Genes involved in the different phases of polycyclic aromatic hydrocarbon (PAH) biodegradation were detected with the functional gene array in the contaminant plume, thus indicating the presence of intrinsic biodegradation potential. However, the low aerobic fluorescein diacetate hydrolysis, the polymerase chain reaction (PCR) amplification of 16S rRNA genes closely similar to sulphate-reducing and denitrifying bacteria and the negligible decrease in contaminant concentrations showed that aerobic PAH biodegradation was limited in the anoxic groundwater. Increased abundance of PAH biodegradation genes was detected by functional gene array in the monitoring well located at the rear end of the biostimulated area, which indicated that air sparging and nutrient infiltration enhanced the intrinsic, aerobic PAH biodegradation. Furthermore, ten times higher naphthalene dioxygenase gene copy numbers were detected by real-time PCR in the biostimulated area, which was in good agreement with the functional gene array data. As a result, functional gene array analysis was demonstrated to provide a potential tool for evaluating the efficiency of the bioremediation treatment for enhancing hydrocarbon biodegradation in field-scale applications. Electronic supplementary material  The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

Method for testing motion analysis laboratory measurement systems

This paper proposes a method for comparing data from accelerometers, optical based 3D motion capture systems, and force platforms (FPs) in the context of spatial and temporal differences. Testing method is based on the motion laboratory accreditation test (MLAT), which can be used to test FP and camera based motion capture components of a motion analysis laboratory. This study extends MLAT to include accelerometer data. Accelerometers were attached to a device similar to the MLAT rod. The elevation of the rod from the plane of the floor is computed and compared with the force platform vector orientation and the rod orientation obtained by optical motion capture system. Orientation of the test device is achieved by forming nonlinear equation group, which describes the components of the measured accelerations. Solution for this equation group is estimated by using the Gauss-Newton method. This expanded MLAT procedure can be used in the laboratory setting were either FP, camera based motion capture, or any other motion capture system is used along with accelerometer measurements.     

Postural control and thigh muscle activity in men with knee osteoarthritis

The aim of this study was to examine the standing balance and the function of vastus medialis (VM) and biceps femoris (BF) muscles with surface electromyography (EMG). Fifty-four subjects with uni- or bilateral knee osteoarthritis (OA) (aged 50–69 years) and 53 age-matched randomly selected clinically and radiologically healthy men participated in this study. Postural control was assessed on a force platform with a bipedal stance with eyes open (EO) and closed (EC) and a monopedal stance with EO. The balance parameters, mean sway velocity, velocity along AP and ML axes, elliptical area, standard deviation of center of pressure, average radial displacement, mean frequency and frequency domain balance parameters and different power spectral density frequency bands were determined. Root mean square (RMS) for EMG amplitude, mean EMG frequency (f_EMG,mean) and median EMG frequency (f_EMG,med) of motor unit activity were calculated from the normalized EMG data. During bipedal stance with EC and EO, there were no significant differences in balance parameters between groups, but during bipedal stance with EO, the RMS in VM was about 56% higher (p < 0.05) in subjects with knee OA than in the control subjects and the values of f_EMG,mean and f_EMG,med were about 48% higher (p < 0.05) in control subjects than subjects with knee OA. It is concluded that subjects with knee OA do not have any standing balance deficit, but they do exhibit increased muscle activity in VM muscle compared to control subjects.     

The metal-binding sites of the zinc-transporting P-type ATPase of Escherichia coli. Lys693 and Asp714 in the seventh and eighth transmembrane segments of ZntA contribute to the coupling of metal binding and ATPase activity

ZntA is a P-type ATPase which transports Zn(2+), Pb(2+) and Cd(2+) out of the cell. Two cysteine-containing motifs, CAAC near the N-terminus and CPC in transmembrane helix 6, are involved in binding of the translocated metal. We have studied these motifs by mutating the cysteines to serines. The roles of two other possible metal-binding residues, K(693) and D(714), in transmembrane helices 7 and 8, were also addressed. The mutation CAAC-->SAAS reduces the ATPase activity by 50%. The SAAS mutant is phosphorylated with ATP almost as efficiently as the wild type. However, its phosphorylation with P(i) is poorer than that of the wild type and its dephosphorylation rate is faster than that of the wild type ATPase. The CPC-->SPS mutant is inactive but residual phosphorylation with ATP could still be observed. The most important findings of this work deal with the prospective metal-binding residues K(693) and D(714): the substitution K693N eliminates the Zn(2+)-stimulated ATPase activity completely, although significant Zn(2+)-dependent phosphorylation by ATP remains. The K693N ATPase is hyperphosphorylated by P(i). ZntA carrying the change D714M has strong metal-independent ATPase activity and is very weakly phosphorylated both by ATP and P(i). In conclusion, K(693) and D(714) are functionally essential and appear to contribute to the metal specificity of ZntA, most probably by being parts of the metal-binding site made up by the CPC motif.     

Enhanced stability of human prion proteins with two disulfide bridges

We compare the folding equilibrium of the globular domain of the human prion protein with two variants of this domain, for which an additional disulfide bond was introduced into the location where it is found in the naturally occurring doppel protein. We find that the unfolding transition midpoint of the variants is shifted toward higher denaturant concentration, indicating that the engineered disulfide bond significantly stabilizes the global protein structure. Our results further reveal that the two-disulfide variant proteins, while possessing the same global fold as the wild-type, display marked differences in their folding pathway-in particular, the absence of a characteristic alpha-helix to beta-sheet transition, which is a fundamental feature associated with misfolding of proteins into amyloid fibrils, especially in the context of prion diseases. These surprising characteristics of disulfide mutant prion proteins have important implications for the understanding of the generic aberrant processes leading to amyloid fibril formation and protein aggregation, as well as providing insight into possible therapeutic strategies.