Reduction of excess sludge production by 3,3′,4′, 5-tetrachlorosalicylanilide in an activated sludge process

The potential of 3,3,4,5-tetrachlorosalicylanilide (TCS) addition to an activated sludge continuous process to reduce excess sludge production by disrupting coupling between anabolism and catabolism was investigated. TCS was chosen as a metabolic uncoupler for continuous test in a lab-scale completely mixed activated sludge process. TCS reduced sludge yield by approximately 30% at a dosage of 40 mg/day. Substrate removal capability was not adversely affected by the presence of TCS, but effluent nitrogen concentration increased during the 60-day continuous operation. Sludge settleability of treated and control samples was qualitatively comparable and not significantly different. Microbial activities in terms of specific oxygen uptake rate were also enhanced, and the microbial population was altered. The results suggest that TCS is an effective chemical uncoupler that reduces sludge yield; process performance was not significantly affected by introduction of the uncoupler.     

Effect of sulfadiazine and trimethoprim on activated sludge performance and microbial community dynamics in laboratory-scale membrane bioreactors and sequencing batch reactors at 8°C

The effect of antibiotics sulfadiazine and trimethoprim on activated sludge operated at 8°C was investigated. Performance and microbial communities of sequencing batch reactors (SBRs) and Membrane Bioreactors (MBRs) were compared before and after the exposure of antibiotics to the synthetic wastewater. The results revealed irreversible negative effect of these antibiotics in environmentally relevant concentrations on nitrifying microbial community of SBR activated sludge. In opposite, MBR sludge demonstrated fast adaptation and more stable performance during the antibiotics exposure. Dynamics of microbial community was greatly affected by presence of antibiotics. Bacteria from classes Betaproteobacteria and Bacteroidetes demonstrated the potential to develop antibiotic resistance in both wastewater treatment systems while Actinobacteria disappeared from all of the reactors after 60 days of antibiotics exposure. Altogether, results showed that operational parameters such as sludge retention time (SRT) and reactor configuration had great effect on microbial community composition of activated sludge and its vulnerability to antibiotics. Operation at long SRT allowed archaea, including ammonium oxidizing species (AOA) such as Nitrososphaera viennensis to grow in MBRs. AOA could have an important role in stable nitrification performance of MBR-activated sludge as a result of tolerance of archaea to antibiotics. © 2018 American Institute of Chemical Engineers Biotechnol. Prog., 35: e2708, 2019     

Can membrane composition traffic toxins? Mycolactone and preferential membrane interactions

Mycolactone is a cytotoxic and immunosuppressive macrolide produced by Mycobacterium ulcerans and the sole causative agent of the neglected tropical skin disease Buruli ulcer. The toxin acts by invading host cells and interacting with intracellular targets to disrupt multiple fundamental cellular processes. Mycolactone’s amphiphilic nature enables strong interactions with lipophilic environments, including cellular membranes; however, the specificity of these interactions and the role of membranes in the toxin’s pathogenicity remain unknown. It is likely that preferential interactions with lipophilic carriers play a key role in the toxin’s distribution in the host, which, if understood, could provide insights to aid in the development of needed diagnostics for Buruli ulcer disease. In this work, molecular dynamics simulations were combined with enhanced free-energy sampling to characterize mycolactone’s association with and permeation through models of the mammalian endoplasmic reticulum (ER) and plasma membranes (PMs). We find that increased order in the PMs not only leads to a different permeation mechanism compared with that in the ER membrane but also an energetic driving force for ER localization. Increased hydration, membrane deformation, and preferential interactions with unsaturated lipid tails stabilize the toxin in the ER membrane, while disruption of lipid packing is a destabilizing force in the PMs.     

Triggers for microbial aggregation in activated sludge?

 Microbial aggregation into good settling sludge is essential for the well-functioning of activated sludge systems treating waste water. Complete aggregation of all the microbial biomass formed has been proven to be difficult to maintain continuously, resulting in wash-out of suspended solids. This review investigates the possibility that environmental signals could constitute triggers for the induction or stimulation of aggregative physiology. Received: 24 August 1995/Accepted: 7 September 1995     

The membrane binding kinetics of full-length PKCα is determined by membrane lipid composition

Protein kinase Cα (PKCα) is activated by its translocation to the membrane. Activity assays show the importance of PIP(2) in determining the specific activity of this enzyme. A FRET stopped flow fluorescence study was carried out to monitor the rapid kinetics of protein binding to model membranes containing POPC/POPS/DOG and eventually PIP(2). The results best fitted a binding mechanism in which protein bound to the membrane following a two-phase mechanism with a first bimolecular reaction followed by a slow unimolecular reaction. In the absence of PIP(2), the rapid protein binding rate was especially dependent on POPS concentration. Formation of the slow high affinity complex during the second phase seems to involve specific interactions with POPS and DOG since it is only sensitive to changes within relatively low concentration ranges of these lipids. Both the association and dissociation rate constants fell in the presence of PIP(2). We propose a model in which PKCα binds to the membranes via a two-step mechanism consisting of the rapid membrane initial recruitment of PKCα driven by interactions with POPS and/or PIP(2) although interactions with DOG are involved too. PKCα searches on the lipid bilayer in two dimensions to establish interactions with its specific ligands.     

Ammonia–methane two-stage anaerobic digestion of dehydrated waste-activated sludge

The study investigated methane production from dehydrated waste-activated sludge (DWAS) with approximately 80% water content under thermophilic conditions. The repeated batch-wise treatment of DWAS using methanogenic sludge unacclimated to high concentrations of ammonia, increased the ammonia production up to 7,600 mg N per kilogram total wet sludge of total ammonia concentration, and stopped the methane production. Investigation revealed that the loading ratio of DWAS for methanogenic sludge influences anaerobic digestion. Methane production significantly decreased and ammonia concentration increased with the increase in loading ratio of DWAS. Since the semicontinuous culture revealed that approximately 50% of organic nitrogen in DWAS converted to ammonia at sludge retention time (SRT) after 4 days at 37 degrees C and 1.33 days at 55 degrees C, the previous stripping of the ammonia produced from DWAS was carried out. The stripping of ammonia increased methane production significantly. This ammonia-methane two-stage anaerobic digestion demonstrated a successful methane production at SRT 20 days in the semicontinuous operation using a laboratory-scale reactor system.     

Biophysics of α-synuclein membrane interactions

Membrane proteins participate in nearly all cellular processes; however, because of experimental limitations, their characterization lags far behind that of soluble proteins. Peripheral membrane proteins are particularly challenging to study because of their inherent propensity to adopt multiple and/or transient conformations in solution and upon membrane association. In this review, we summarize useful biophysical techniques for the study of peripheral membrane proteins and their application in the characterization of the membrane interactions of the natively unfolded and Parkinson's disease (PD) related protein, α-synuclein (α-syn). We give particular focus to studies that have led to the current understanding of membrane-bound α-syn structure and the elucidation of specific membrane properties that affect α-syn-membrane binding. Finally, we discuss biophysical evidence supporting a key role for membranes and α-syn in PD pathogenesis. This article is part of a Special Issue entitled: Membrane protein structure and function.     

Did membrane electrochemistry precede translation?

The discovery of RNA-based enzymes, such as ribonuclease-P, has stimulated new interest in the idea that catalytic functions of RNA preceded the use of coded enzymes during an era loosely termed the RNA world. This paper examines various lines of evidence which support the idea that electrochemical processes associated with the membrane may have preceded the development of coded protein enzymes and may have provided a basis for the phosphorylation energy of the RNA world.     

Vesicle-associated membrane protein 2 mediates trafficking of α5β1 integrin to the plasma membrane

Integrins are major receptors for cell adhesion to the extracellular matrix (ECM). As transmembrane proteins, the levels of integrins at the plasma membrane or the cell surface are ultimately determined by the balance between two vesicle trafficking events: endocytosis of integrins at the plasma membrane and exocytosis of the vesicles that transport integrins. Here, we report that vesicle-associated membrane protein 2 (VAMP2), a SNARE protein that mediates vesicle fusion with the plasma membrane, is involved in the trafficking of α5β1 integrin. VAMP2 was present on vesicles containing endocytosed β1 integrin. Small interfering RNA (siRNA) silencing of VAMP2 markedly reduced cell surface α5β1 and inhibited cell adhesion and chemotactic migration to fibronectin, the ECM ligand of α5β1, without altering cell surface expression of α2β1 integrin or α3β1 integrin. By contrast, silencing of VAMP8, another SNARE protein, had no effect on cell surface expression of the integrins or cell adhesion to fibronectin. In addition, VAMP2-mediated trafficking is involved in cell adhesion to collagen but not to laminin. Consistent with disruption of integrin functions in cell proliferation and survival, VAMP2 silencing diminished proliferation and triggered apoptosis. Collectively, these data indicate that VAMP2 mediates the trafficking of α5β1 integrin to the plasma membrane and VAMP2-dependent integrin trafficking is critical in cell adhesion, migration and survival.     

The major basement membrane components localize to the chondrocyte pericellular matrix--a cartilage basement membrane equivalent?

In this study, we demonstrate that articular cartilage chondrocytes are surrounded by the defining basement membrane proteins laminin, collagen type IV, nidogen and perlecan, and suggest that these form the functional equivalent of a basement membrane. We found by real-time PCR that mouse chondrocytes express these four cardinal components of basement membranes and demonstrated by immunohistochemistry that the proteins are present in bovine and mouse cartilage tissues and are deposited in a thin pericellular structure. Immunoelectron microscopy confirmed high laminin concentration in the pericellular matrix. In cartilage from newborn mice, basement membrane components are widespread in the territorial and interterritorial matrix, while in mature cartilage of adult mice the basement membrane components are localized mainly to a narrow pericellular zone. With progression into old age, this layer becomes less distinct, especially in areas of obvious mechanical attrition. Interestingly, individual laminin subunits were located in different zones of the cartilage, with laminin alpha1 showing preferential localization around a select population of superficial layer chondrocytes. We propose that the chondrocyte, like several other cell types of mesenchymal origin, is surrounded by the functional equivalent of a basement membrane. This structure is presumably involved in maintaining chondrocyte phenotype and viability and may well allow a new understanding of cartilage development and provide clues to the progression of degenerative joint disorders.     

Simultaneous sewage sludge digestion and metal leaching — effect of temperature

The effect of temperature on the simultaneous sewage sludge digestion and metal leaching process was studied in laboratory bioreactors of 20 l working volume. The results thus obtained showed that the process can be employed efficiently for metal solubilization, elimination of indicator microorganisms and sewage sludge stabilization at temperatures between 10°C and 30°C. Rates of pH reduction, sulfur oxidation, growth of thiobacilli, elimination of indicator microorganisms and solids degradation were found to decrease with temperature. Low metal solubilization efficiency was observed at 10°C; however, metals were solubilized to below the recommended level. The solubilization of organic matter and nutritive elements (N, P and K) was not significantly affected by the variation in temperature. The fertilizer value of sludge after leaching and digestion did not change significantly and remained the same irrespective of temperature. *** DIRECT SUPPORT *** AG903078 00005     

Incorporation of β-sitosterol into mitochondrial membrane enhances mitochondrial function by promoting inner mitochondrial membrane fluidity

Recent findings suggest that mitochondrial membrane fluidity could influence mitochondrial energy metabolism. β-sitosterol (BS) is a common plant sterol that is prevalent in plant oils, nuts, cereals and plant food products. Its chemical structure is very similar to that of cholesterol. As a cholesterol analog, BS is highly lipid soluble and largely resides in the membranes of cells or organelles where it may have an influence on the membrane fluidity. The present study reports that, with the cholesterol chelator 2-hydroxypropyl-β-cyclodextrin (HPβCD) as its carrier, BS is able to increase the fluidity of the inner mitochondrial membrane (IMM) without affecting the fluidity of the outer mitochondrial membrane (OMM), and consequently to increase the mitochondrial membrane potential (?Ψm) and mitochondrial ATP content. It has been previously proposed that a therapeutical boost in adenosine triphosphate (ATP) levels in mitochondria may be beneficial for neurodegenerative diseases such as Alzheimer’s disease (AD). Given that dietary administration of plant sterols could increase brain BS concentrations, these results may provide a better understanding of the beneficial effects of plant sterol-enriched nutrients on neurodegenerative diseases such as AD.     

Sticholysin I–membrane interaction: An interplay between the presence of sphingomyelin and membrane fluidity

Sticholysin I (St I) is a pore-forming toxin (PFT) produced by the Caribbean Sea anemone Stichodactyla helianthus belonging to the actinoporin protein family, a unique class of eukaryotic PFT exclusively found in sea anemones. As for actinoporins, it has been proposed that the presence of sphingomyelin (SM) and the coexistence of lipid phases increase binding to the target membrane. However, little is known about the role of membrane structure and dynamics (phase state, fluidity, presence of lipid domains) on actinoporins' activity or which regions of the membrane are the most favorable platforms for protein insertion. To gain insight into the role of SM on the interaction of St I to lipid membranes we studied their binding to monolayers of phosphatidylcholine (PC) and SM in different proportions. Additionally, the effect of acyl chain length and unsaturation, two features related to membrane fluidity, was evaluated on St I binding to monolayers. This study revealed that St I binds and penetrates preferentially and with a faster kinetic to liquid-expanded films with high lateral mobility and moderately enriched in SM. A high content of SM induces a lower lateral diffusion and/or liquid-condensed phases, which hinder St I binding and penetration to the lipid monolayer. Furthermore, the presence of lipid domain borders does not appear as an important factor for St I binding to the lipid monolayer.     

Intracellular membrane fusion: SNAREs only?

The past two years have seen vigorous attempts to elucidate the mechanism driving intracellular membrane fusion. Much attention was focused on the role of SNARE complexes. Their crystal structure was solved and fusion was reconstituted using proteoliposomes with purified SNAREs suggesting them to be the minimal machinery for fusion. Work on physiological membranes, however, points in another direction and has spurred a hot debate on the function of SNAREs.     

Is arginine charged in a membrane?

“Charged” amino acids play countless important roles in protein structure and function. Yet when these side chains come into contact with membranes we do not fully understand their behavior. This is highlighted by a recent model of voltage-gated ion channel activity and translocon-based experiments that suggest small penalties to expose these side chains to lipids, opposing the prevailing view in membrane biophysics. Here we employ a side chain analog as well as a transmembrane helix model to determine the free energy as a function of protonation state and position for a lipid-exposed arginine (Arg) residue across a membrane. We observe high free energy barriers for both the charged and neutral states. Due to the stabilizing influence of membrane deformations for the protonated form, the Arg side chain experiences a pK_a shift of ≤4.5 units and remains mostly protonated. The cost for exposing Arg to lipid hydrocarbon is prohibitively high with implications for many membrane translocating processes and the activation mechanisms of voltage-gated ion channels.     

Membrane phosphoinositides and protein–membrane interactions

Proteins with polybasic clusters bind to negatively charged phosphoinositides at the cell membrane. In this review, I have briefly discussed the types of phosphoinositides naturally found on membrane surfaces and how they recruit protein complexes for carrying out the process of signal transduction. A large number of researchers from around the world are now focusing their attention on protein–membrane binding, as these interactions have started to offer us a much better insight into the process of cell signaling. The main areas discussed in this brief review article include the phosphoinositide binding specificities of proteins and the role of their lipid binding in signaling processes downstream of membrane recruitment.     

What drives membrane fusion in eukaryotes?

The fusion of biological membranes is the terminal step of all vesicular trafficking reactions in eukaryotic cells. Therefore, this fusion is fundamental for the transfer of proteins and lipids between different compartments, for exocytosis and for the structural integrity of organelles. In the past decade, many parts of the molecular machinery involved in fusion have been uncovered. Although the mechanisms responsible for mutual recognition and binding of membranes inside eukaryotes are becoming reasonably well known, there is considerable uncertainty as to what causes the actual merging of the lipid bilayer. Two classes of mechanisms have been proposed. Proximity models postulate that very close apposition of membranes suffices to induce fusion. By contrast, pore models propose that continuous proteinaceous pores between apposed membranes could be the basis for fusion.