Journal Of Biological Chemistry Research Articles

Abstract SY20-02: Insights into mechanisms of breast cancer metastasis

Abstract SY20-02: Insights into mechanisms of breast cancer metastasis

Voltammetry Behavior of Carbon Paste Electrodes with Native and Chemical Modified Porphyrans

Porphyra species are important edible red algae abundantly cultivated; they are commonly known as ‘nori’ and used to prepare sushi. P. tenera and P. yezoensis are the species cultured and processed into this sheet type dried food. Porphyran is a linear sulfated water soluble polysaccharide that can be obtained from Porphyra species1,2. It is constituted by units of (6-O-methyl)-β-D-galactose, 3,6-anhydro-α-L-galactose and α-L-galactose 6-sulfate1,2. Carbon paste electrodes (CPE) containing natural ionic polysaccharides (pectic and alginic acid and heparin) have been studied3 and biosensors like these are generally developed for the detection of analytes particularly in aqueous phase4. The present work describes the voltammetric behavior of porphyran-modified CPE obtained before and after chemical desulfation of the polysaccharides. Porphyran was obtained from dried milled nori by water extraction (1.5 %w/v, for 16 h, at room temperature) and precipitation with ethanol (three volumes). The native fraction (PN) was purified by redissolution in water, dialysis and lyophilization. The desulfated porphyrans fractions were obtained by solvolysis5 (PD) and alkaline treatment6 (PA) from the native fraction. The polysaccharides structures were confirmed by 13C NMR analysis. The carbon paste was prepared by hand-mixing the porphyran with graphite powder (100 mg) and mineral oil (30 µL). Electrodes containing 10, 20, 40, 60, 80 and 100 µg of the native (EN) and chemically modified porphyran (ED and EA) were studied. The carbon paste was packed into the hole of the electrode body (working electrode) and an Ag/AgCl (reference electrode) and a platinum wire (auxiliary electrode) completed the three-electrode system in the measurement cell. Cyclic voltammograms (CV) were recorded in the potential range of -0.6 – 1.0 V (PBS buffer 0.15 mol L-1, pH 6.5) with a scan rate of 50 mV s-1 using K3Fe(CN) 6 /K4Fe(CN) 6 0.1 mol L-1 as redox reference solution. The measurements were performed using a PalmSens potentiostat (Palm Instruments BV). The NMR spectra obtained for PN, PD and PA confirmed the porphyran structural pattern as they also showed that the chemical treatments produced desulfated polysaccharides. Especially for PA fraction, the NMR spectrum indicated an increase in the 3,6-anhydrogalactose content as a result of cyclization of the galactose-6-sulfate, as expected by the alkaline treatment7. The CVs was used to characterize the porphyran-modified CPEs and the voltammograms indicated an increase in the cathodic and anodic current peaks in the same potential range for all electrodes as compared to unmodified CPE. Despite this, the increase in current values did not correlate directly with the porphyran concentration used to modify the electrodes, so the CPEs with 10, 20 and 40 µg showed the most intense anodic peaks for EN (3,648.81 µA), EA (3,838.67 µA) and ED (4,452.00 µA), respectively (Figure 1). The higher values of redox current peaks were found for the electrode ED at all porphyrans concentrations studied. This behavior can be attributed to electrostatic repulsion between the K3Fe(CN) 6 /K4Fe(CN) 6 and the ionic polysaccharide. As the porphyran PD has a lower number of sulfate groups, the electrode ED may have a better electrochemical response. In addition, the peak currents increased and the cathodic and anodic peak potential exhibited a small shift along with the increase of scan rate for EN (10ug), EA (20ug) and ED (40ug). At the same time, the anodic peak current increased linearly with the square root of the scan rate (correlation coefficient of 0.9711, 0.9975 and 0.9943 for EN, EA and ED respectively). This phenomenon suggested that the redox process was diffusion-controlled. In conclusion, the CVs demonstrated that the electron transfer on the porphyran-modified CPEs surface, especially on ED, are more efficient, conducting higher current and offering lower resistivity.1- JIANG, Z.; HAMA, Y.; YAMAGUCHI, K.; ODA, T. J. Biochem., v. 151, p. 65, 2012.2- ISHIHARA, K.; OYAMADA, C.; MATSUSHIMA, R., et al. Biosci. Biotechnol. Biochem., v. 69, p. 1824, 2005.3- WANG, J.; TAHA, Z.; NASER, N. Talanta, v. 38, p. 81, 1991. 4- MOHSENI, G.; NEGAHDARY, M.; FARAMARZI, H.; et al. Int. J. Electrochem. Sci., v. 7, p. 12098, 2012. 5- CIANCIA, M.; NOSEDA, M.D.; MATULEWICZ, M.C.; CEREZO, A.S. Carbohydrate Polymers, v. 20, p. 95, 1993.6- NAGASAWA, K.; INOUE, Y.; TOKUYASU, T. Journal of Biological Chemistry, v. 86, p. 1323, 1979.Figure 1: Cyclic voltammogram of EN (10 µg), EA (20 µg) and ED (40 µg), in PBS buffer 0.15 mol L-1, pH 6.5, potential range of -0.6 – 1.0 V, using K3Fe(CN) 6 /K4Fe(CN) 6 0.1 mol L-1 as redox reference solution.

Brain mitochondria from DJ-1 knockout mice show increased respiration-dependent hydrogen peroxide consumption

Mutations in the DJ-1 gene have been shown to cause a rare autosomal-recessive genetic form of Parkinson’s disease (PD). The function of DJ-1 and its role in PD development has been linked to multiple pathways, however its exact role in the development of PD has remained elusive. It is thought that DJ-1 may play a role in regulating reactive oxygen species (ROS) formation and overall oxidative stress in cells through directly scavenging ROS itself, or through the regulation of ROS scavenging systems such as glutathione (GSH) or thioredoxin (Trx) or ROS producing complexes such as complex I of the electron transport chain. Previous work in this laboratory has demonstrated that isolated brain mitochondria consume H2O2 predominantly by the Trx/Thioredoxin Reductase (TrxR)/Peroxiredoxin (Prx) system in a respiration dependent manner (Drechsel et al., Journal of Biological Chemistry, 2010). Therefore we wanted to determine if mitochondrial H2O2 consumption was altered in brains from DJ-1 deficient mice (DJ-1−/−). Surprisingly, DJ-1−/− mice showed an increase in mitochondrial respiration-dependent H2O2 consumption compared to controls. To determine the basis of the increased H2O2 consumption in DJ1−/− mice, the activities of Trx, Thioredoxin Reductase (TrxR), GSH, glutathione disulfide (GSSG) and glutathione reductase (GR) were measured. Compared to control mice, brains from DJ-1−/− mice showed an increase in (1) mitochondrial Trx activity, (2) GSH and GSSG levels and (3) mitochondrial glutaredoxin (GRX) activity. Brains from DJ-1−/− mice showed a decrease in mitochondrial GR activity compared to controls. The increase in the enzymatic activities of mitochondrial Trx and total GSH levels may account for the increased H2O2 consumption observed in the brain mitochondria in DJ-1−/− mice perhaps as an adaptive response to chronic DJ-1 deficiency.

Neutralizing Vacuoles of C. albicans Can Tame Its Virulence

When the vacuoles of Candida albicans cells are made less acidic, this fungal pathogen becomes less virulent and potentially more manageable as an infectious agent, according to Rajini Rao of Johns Hopkins University School of Medicine in Baltimore, Md., and her collaborators. Details appeared July 24, 2013 in the Journal of Biological Chemistry (doi:10.1074/jbc.M113.494815).

Podocyte-specific deletion of NDST1, a key enzyme in the sulfation of heparan sulfate glycosaminoglycans, leads to abnormalities in podocyte organization in vivo

Heparan sulfate proteoglycans have been shown to modulate podocyte adhesion to- and pedicel organization on- the glomerular basement membrane. Recent studies showed that foot process effacement developed in a mutant mouse model whose podocytes were unable to assemble heparan sulfate glycosaminoglycan chains. This study, a further refinement, explored the role of heparan N-sulfation on podocyte behavior. A novel mutant mouse (Ndst1-/-) was developed, having podocyte-specific deletion of NDST1, the enzyme responsible for N-sulfation of heparan sulfate chains. Podocytes having this mutation had foot process effacement and abnormal adhesion to Bowman's capsule. Although glomerular hypertrophy did develop in the kidneys of mutant animals, mesangial expansion was not seen. The lack of heparan N-sulfation did not affect the expression of agrin or perlecan proteoglycan core proteins. Loss of N-sulfation did not result in significant proteinuria, but the increase in the albumin/creatinine ratio was coincident with the development of the enlarged lysosomes in the proximal tubules. Thus, although the renal phenotype of the Ndst1-/- mouse is mild, the data show that heparan chain N-sulfation plays a key role in podocyte organization.

Comment onTimely deposition of macromolecular structures is necessary for peer reviewby Joostenet al.(2013)

The wwPDB responds to the article by Joostenet al.[(2013),Acta Cryst.D69, 2293–2295].

A tight junction between E-Cadherin and the prostate tumor suppressor SPDEF

A tight junction between E-Cadherin and the prostate tumor suppressor SPDEF

Abstract 2325: Identification of RACK1 as a novel regulatory subunit of PP2A.

Abstract Protein Phosphatase 2A (PP2A) is a major serine/threonine phosphatase in cells. It consists of a catalytic (C), structural (A) and regulatory/variable B-type subunits. PP2A plays a critical role as a tumour suppressor. Changes in the activity of PP2A have a direct role in maintaining the transformed phenotype in cancer. However, the mechanism of action of PP2A during this process is poorly understood. Understanding is limited by the complex nature of PP2A holoenzyme assembly, and because it acts through a wide variety of signalling pathways. The regulatory B-type subunits have a critical role in targeting PP2A to specific areas of the cell and also in regulating the overall activity and substrate specificity of PP2A. To date, at least 15 known PP2A B-type subunits have been identified but their detection is difficult because of the transient nature of their interaction with the PP2A holoenzyme. The scaffolding protein RACK1 is a PP2A interacting protein and we have published a mechanism by which PP2A regulates IGF-IR and adhesion signalling through RACK1 [1, 2]. Our hypothesis is that RACK1 plays a critical role in determining how PP2A functions in signalling pathways and may function as a regulatory subunit of PP2A. We predict it does this by modulating the activity of PP2A and regulating its distribution to a specific subset of targets. We examined how the PP2A holoenzyme is assembled in MCF-7 cells. We have found that both the catalytic subunit and structural subunit of PP2A (PP2A C and PP2A A respectively) are assembled on the RACK1 scaffold in response to IGF-I. Our findings indicate that both the C subunit and the A subunit of PP2A are released from RACK1 in response to IGF-I and reassembled after 30 minutes IGF-I stimulation. We used immobilized peptide arrays representing the entire PP2A-catalytic protein to identify amino acids on the catalytic subunit of PP2A that are required for the binding of RACK1. Analysis of these residues revealed the binding sites for RACK1 on PP2A and supports their role in mediating the interaction with RACK1. When cells are transfected with specific mutations of PP2A C that disrupt the association with RACK1, cells show decreased PP2A activity and deregulated cell adhesion and migration suggesting that RACK1 has an essential role to play in the overall activity and substrate specificity of PP2A. This data supports our hypothesis that RACK1 functions as a B-type subunit of PP2A which is required for the regulation of PP2A activity. Our data also indicates that RACK1 targets PP2A to specific locations to promote IGF-I-mediated cell migration and proliferation in tumour cells. 1. Kiely, P.A., et al. Molecular and cellular biology, 2006. 26(11): p. 4041-4051. 2. Kiely, P.A., et al. Journal of Biological Chemistry, 2008. 283(34): p. 22952-22961. Citation Format: Maeve L. Kiely, Rosemary O'Connor, Patrick A. Kiely. Identification of RACK1 as a novel regulatory subunit of PP2A. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 2325. doi:10.1158/1538-7445.AM2013-2325

Freely diffusing versus adsorbed protein: Which better mimics the cellular state of a redox protein?

Dynamic electrochemistry of adsorbed proteins, often termed protein film electrochemistry (PFE), is widely used for the characterisation of redox proteins. The method provides a powerful alternative to spectroscopic studies that interrogate protein solutions. The reduction potential and electron stoichiometry of redox couples can be defined. The rates of catalytic redox transformations can also be quantified. Often it is considered that the behaviour of the adsorbed protein should match that displayed in solution studies if it is to be relevant to understanding the biological role of the protein. However, the environment of the protein in PFE is fundamentally different from that when it is freely diffusing in solution. As a consequence different behaviours may be expected. This raises the question, which approach is more relevant when aiming to provide insight into the cellular role of the protein? We consider this question here taking as an example the properties of a penta-heme cytochrome NrfB from Escherichia coli. The redox properties of NrfB containing solutions were presented previously (Clarke et al., Journal of Biological Chemistry (2004)). Here we present PFE of NrfB adsorbed on graphite and optically transparent mesoporous, nanocrystalline SnO2 electrodes.

Inositol, Glycogen, Insulin, and Six Nobelists

My reflections cover my scientific work over a sixty-year period. At the University of Michigan, I was initially interested in embryology. In medical school in the army, I became entranced with biochemistry and DeWitt Stetten, who introduced me to Carl Cori. Carl along with Gerty Cori and Earl Sutherland became my scientific mentors. After medical school and a nine-month medical internship, I re-entered the army and was assigned to Edgewood Arsenal in Maryland, where I established the NAD requirement for brain pyruvate dehydrogenase to oxidize pyruvate and published my first Journal of Biological Chemistry (JBC) paper. At the University of Illinois, I obtained an M.S. degree in chemistry and was influenced by Will Rose and Herb Carter. At Washington University in St. Louis, I worked with Gerty and Carl Cori on glycogen metabolism and obtained my Ph.D. degree. I also worked with Bill Daughaday on the increased urinary excretion of myo-inositol in diabetic subjects and in rats preloaded with glucose. I was offered an instructor's appointment in biochemistry at Washington University, but I decided to accept an assistant professor's appointment in the Division of Biochemistry of the Chemistry Department at the University of Illinois. I spent four years at Illinois working on intestinal starch digestion with particular reference to glycosidases hydrolyzing the α1,6-branch point linkages. I was recruited by Earl Sutherland to join his Department of Pharmacology as an associate professor at Western Reserve University in Cleveland. Here, Carlos Villar-Palasi, my first postdoctoral fellow, and I began our insulin studies, which led to the discovery of two covalent forms of glycogen synthase controlled by insulin and epinephrine. These continued at the University of Minnesota, where I held an endowed chair in biochemistry. Here, with students and postdoctoral fellows, we further defined the two forms of glycogen synthase and worked on the possible role of cAMP in insulin action. Moving to the University of Virginia as Chair of Pharmacology, I recruited two future Nobelists, Al Gilman and Ferid (Fred) Murad, with Ted Rall, their past mentor for their Ph.D. degrees, as well as other faculty who would become departmental chairs. Our early studies on glycogen and inositol became united with studies on insulin action. We isolated a novel inositol glycan pseudo-disaccharide insulin second messenger containing a rare inositol, d-chiro-inositol (DCI). Its structure was determined as galactosamine β1,4-pinitol, where pinitol is the 3-O-methyl ether of DCI. It was chemically synthesized. We and others found that insulin resistance in several human diseases is related to an inositol imbalance consisting of a deficit of chiro-inositol and an excess of myo-inositol, an unfolding story now under way involving the myo-inositol to chiro-inositol epimerase. I am blessed to have had such superlative scientific mentors, a productive group of students and postdoctoral fellows, a loving wife of sixty-five years, three remarkable sons, and eight wonderful grandchildren. I am working full time on insulin, which has entranced me for a long time.

Assessing the Safety of a Replacement Chemical: Nongenomic Activity of Bisphenol S

Suspected links between bisphenol A (BPA) and adverse human health effects have spurred a search for replacement chemicals for use in common applications such as polycarbonate plastics, food can linings, and thermal papers. Bisphenol S (BPS) has been adopted for some uses. A new study now shows that low doses of BPS can disrupt nongenomic signaling pathways in cultured pituitary cells [EHP 121(3):352–358; Vinas and Watson]. Nearly 93% of U.S. residents over age 6 carry measurable amounts of BPA in their bodies. The compound is suspected to contribute to the development of diseases such as diabetes, asthma, and cancer, and animal studies have shown it adversely affects reproduction. In contrast to genomic regulation, which involves receptors in the cell nucleus, nongenomic regulation involves receptors in the cell membrane. Since BPA can interact with cell-membrane estrogen receptors, researchers explored whether structurally similar BPS might likewise influence nongenomic pathways. Treatments included various low doses of BPS alone and paired with estradiol, an endogenous estrogen. Measured responses included activation of ERK and JNK, mitogen-activated protein kinases involved in cell growth, proliferation, and apoptosis (programmed cell death); activation of caspases 8 and 9, enzymes that also are involved in apoptosis; and release of prolactin, a hormone that helps regulate hundreds of biological functions, including metabolism, reproduction, and lactation. BPS has been adopted as a replacement for BPA in applications such as thermal receipt paper. BPS appeared to interact predominantly with the cell-membrane estrogen receptor ER〈. Estradiol and BPS each increased cell proliferation, but following exposure to both compounds there were fewer cells than in control cultures. This can result from decreased proliferation or increased cell death, which is why the investigators also looked at caspase 8 and caspase 9. The former was activated by BPS alone and when combined with estradiol, but caspase 9 was less affected, consistent with an increase in apoptosis. The lowest concentrations of BPS triggered the strongest ERK response; however, ERK activation was not as strong in the presence of both compounds. BPS alone did not activate JNK, but in combination with estradiol it increased JNK activation more than the estrogen alone. The quality and timing of ERK and JNK responses to activation by estradiol were often altered by BPS. By itself, BPS did not affect prolactin secretion, but it significantly suppressed estradiol-induced secretion at most concentrations. Based on these results, low concentrations of BPS appear to affect nongenomic signaling in estrogen-responsive cells, with potential consequences for cell function. The results emphasize the need to screen compounds for estrogenic activity prior to their use in manufacturing.

Muscle dysfunction in COPD

Although it has been well characterized that COPD muscle dysfunction is the result of the complex interaction between systemic and local factors, the etiology of muscle dysfunction remains to be fully identified. Interestingly, while hyperinflation and an increased work of breathing appear to be the main contributing events to respiratory muscle dysfunction, deconditioning seems to play a major role in the dysfunction of peripheral muscles in COPD. Other factors such as cigarette smoke, nutritional abnormalities, exacerbations, drugs, hypoxia, hypercapnia, comorbidities, and physical activity also influence muscle mass and function in COPD patients (1-3; 5; 9; 15). Besides, derangements of key molecular and cellular processes such as redox imbalance, mitochondrial dysfunction, enhanced protein catabolism and reduced protein anabolism, structural alterations, and systemic inflammation also modify muscle phenotype and function in COPD patients.

Data Submission and Quality in Microarray-Based MicroRNA Profiling

Public sharing of scientific data has assumed greater importance in the omics era. Transparency is necessary for confirmation and validation, and multiple examiners aid in extracting maximal value from large data sets. Accordingly, database submission and provision of the Minimum Information About a Microarray Experiment (MIAME)(3) are required by most journals as a prerequisite for review or acceptance. In this study, the level of data submission and MIAME compliance was reviewed for 127 articles that included microarray-based microRNA (miRNA) profiling and were published from July 2011 through April 2012 in the journals that published the largest number of such articles--PLOS ONE, the Journal of Biological Chemistry, Blood, and Oncogene--along with articles from 9 other journals, including Clinical Chemistry, that published smaller numbers of array-based articles. Overall, data submission was reported at publication for <40% of all articles, and almost 75% of articles were MIAME noncompliant. On average, articles that included full data submission scored significantly higher on a quality metric than articles with limited or no data submission, and studies with adequate description of methods disproportionately included larger numbers of experimental repeats. Finally, for several articles that were not MIAME compliant, data reanalysis revealed less than complete support for the published conclusions, in 1 case leading to retraction. These findings buttress the hypothesis that reluctance to share data is associated with low study quality and suggest that most miRNA array investigations are underpowered and/or potentially compromised by a lack of appropriate reporting and data submission.

Editorial introductions

Editorial introductions

Unexpected Role for Heme Protein in Archaeon M. acetivorans

The protein MA4561 from the archaeon Methanosarcina acetivorans is not the phytochrome-like photoreceptor that it was first thought to be, but is instead a heme-binding protein that likely acts as a redox sensor, according to Nicole Frankenberg-Dinkel at the Ruhr-Universitat in Bochum, Germany, and her collaborators. While some bacteria use heme-containing proteins as redox sensors, M. acetivorans is the first archaeon known to do so, she says. Details appeared May 9, 2013 in the Journal of Biological Chemistry (doi: 10.1074/jbc.M113.476267 jbc.M113.476267).

Probing the Stoichiometry and Geometry of M3 Acetylcholine Receptors at the Plasma Membrane

G-protein-coupled receptors (GPCRs) are the largest family of transmembrane proteins in nature. GPCRs cascade signals from outer environment into cells and hence, are the target of more than 60% of modern clinical drugs. Determination of oligomerization of GPCRs is subject to significant controversy in the literature. We have investigated the quaternary structure of human muscarinic acetylcholine receptor type 3 (hM3) in living cells using Forster Resonance Energy Transfer (FRET) and two-photon excitation in an optical micro-spectroscopic set-up [Raicu et al, Nature Photonics, 2009]. The wild-type form of the hM3 receptor was fused to Cerulean while its mutated form, activated solely by a synthetic ligand (RASSL), was fused to Citrine and expressed constitutively in Flp-InTM T-RExTM 293 cells [Alvarez-Curto et al, Journal of Biological Chemistry, 2010]. When WT-hM3-Citrine and RASSL-hM3-Cerulean were co-expressed in same cells, excited donor (Cerulean), transferred its energy to nearby acceptors (Citrine) through FRET. Apparent FRET efficiency (Eapp) distribution maps were obtained for the imaged section of the cells by unmixing acquired spectrally resolved images using elementary donor and acceptor spectra. By selecting pixels in the Eapp image corresponding to the plasma membrane, a FRET efficiency histogram was obtained displaying the number of pixels in a particular range of Eapp values versus the corresponding Eapp value. The Eapp histograms were analyzed using a FRET theory [V. Raicu, Journal of Biological Physics, 2007], predicting various numbers and positions of peaks in the histograms for various sizes and geometry of oligomers. Eapp histograms of all cells were best fitted by a rhombus tetramer model. By also simulating the amplitudes of the peaks, we determined that hM3 receptors form both dimers and rhombus tetramers at the plasma membrane, and that their proportion remained largely unaffected by CNO binding.

THE HUMBLE BEGINNINGS OF THE STUDY OF NITROGEN EXCRETION IN FISH

> Tamara Rodela discusses Homer William Smith's classic paper ‘The excretion of ammonia and urea by the gills of fish’, published in the Journal of Biological Chemistry in 1929. Homer William Smith (1895–1962) was best known for his work as a renal physiologist. His extensive research on the

Rio+20: Challenges of Development for Health and Sustainability

Vol. 120, No. 9 EditorialOpen AccessRio+20: Challenges of Development for Health and Sustainability Maria Cecília de Souza Minayo Maria Cecília de Souza Minayo Search for more papers by this author Published:1 September 2012https://doi.org/10.1289/ehp.1205814Cited by:2AboutSectionsPDF ToolsDownload CitationsTrack Citations ShareShare onFacebookTwitterLinked InReddit The United Nations Conference on Sustainable Development (Rio+20; United Nations 2011) took place in Rio de Janeiro, Brazil, on 20–22 June 2012. This conference brought together politicians, leaders from social movements, and business representatives from around the world. One goal of the conference was to renew commitment to sustainable development and ensure the promotion of an economically, socially, and environmentally sustainable future for the planet for present and future generations.The Future We Want (United Nations 2012), the zero draft of the outcome document from Rio+20, listed seven priority areas: job creation, food security, water, energy, sustainable cities, oceans, and natural disasters. Unfortunately, as pointed out in a recent editorial from The Lancet (2012), human health was hardly mentioned. This is in contrast to Rio-92, where health was viewed as a major theme in Chapter 6 of Agenda 21 (United Nations 1992). The Lancet (2012) emphasized the need to ensure that the health agenda of the past decade is completed and that new and emerging concerns that could affect human health, such as noncommunicable diseases and climate change, are considered.In the past few years, there have been many positive changes in human health and the environment. To demonstrate this point, the June 2012 issue of Ciência & Saúde Coletiva (Vol. 17, No. 6; http://www.scielo.br/scielo.php?script=sci_issuetoc&pid=1413-812320120006&lng=en&nrm=iso) was devoted to the changes in human health and the environment in Brazil. Articles in the issue indicated decreases in infant mortality, increases in life expectancy, and decreases in maternal mortality. In addition to these changes, deforestation and burning of forests has declined; the use of technologies that pollute has declined; inspection and surveillance mechanisms relevant to food, water, and air quality have been created; agroecology has expanded; forests have been replanted; and environmental preservations areas have been established. However, Brazil still faces immense environmental liabilities that could affect human health, including the increasing use of agricultural monoculture, continued high-volume use of agricultural pesticides and fertilizers, adverse health impacts from large-scale enterprises (e.g., construction of hydroelectric power plants, sugar and alcohol production, soybean agriculture, raising of livestock), the lack of basic sanitation and solid waste treatment for much of the Brazilian population, increased deforestation and forest burning, the persistence of asbestos and heavy metals, and the unequal distribution of water or lack of clean drinking water.Many of these health-related issues are not unique to Brazil. In fact, many developed and developing countries face similar environmental challenges that are affecting or could affect human health. Although it is important to promote sustainable development in the future, there is concern about the virtual absence of health-related goals from Rio+20. Ciência & Saúde Coletiva endorses the call by The Lancet (2012) to move toward integrating health and sustainability goals in any future agenda developed by the United Nations.The author declares she has no actual or potential competing financial interests.ReferencesThe Lancet. 2012. Global health in 2012: development to sustainability [Editorial].Lancet 379(9812):19322265616. Crossref, Medline, Google ScholarUnited Nations. 1992. Agenda 21. Protecting & Promoting Human Health.Available: http://www.un.org/esa/dsd/agenda21/res_agenda21_06.shtml [accessed 7 August 2012]. Google ScholarUnited Nations. 2011. Rio+20: United Nations Conference on Sustainable Development.Available: http://www.uncsd2012.org/ [accessed 7 August 2012]. Google ScholarUnited Nations. 2012. The Future We Want—Zero Draft of the Outcome Document.Available: http://www.uncsd2012.org/rio20/index.php?page=view&type=12&nr=324&menu=23 [accessed 7 August 2012]. Google ScholarFiguresReferencesRelatedDetailsCited By Quintana C, Olea P, Abdallah P and Quintana A (2016) Port environmental management: Innovations in a Brazilian public port, RAI Revista de Administração e Inovação, 10.1016/j.rai.2016.09.001, 13:4, (261-273), Online publication date: 1-Oct-2016. Filograna R, Godena V, Sanchez-Martinez A, Ferrari E, Casella L, Beltramini M, Bubacco L, Whitworth A and Bisaglia M (2016) Superoxide Dismutase (SOD)-mimetic M40403 Is Protective in Cell and Fly Models of Paraquat Toxicity, Journal of Biological Chemistry, 10.1074/jbc.M115.708057, 291:17, (9257-9267), Online publication date: 22-Apr-2016. Vol. 120, No. 9 September 2012Metrics About Article Metrics Publication History Originally published31 August 2012Published in print1 September 2012 Financial disclosuresPDF download License information EHP is an open-access journal published with support from the National Institute of Environmental Health Sciences, National Institutes of Health. All content is public domain unless otherwise noted. Note to readers with disabilities EHP strives to ensure that all journal content is accessible to all readers. However, some figures and Supplemental Material published in EHP articles may not conform to 508 standards due to the complexity of the information being presented. If you need assistance accessing journal content, please contact [email protected]. Our staff will work with you to assess and meet your accessibility needs within 3 working days.

Two ATPases

In this article, I reflect on research on two ATPases. The first is F(1)F(0)-ATPase, also known as ATP synthase. It is the terminal enzyme in oxidative phosphorylation and famous as a nanomotor. Early work on mitochondrial enzyme involved purification in large amount, followed by deduction of subunit composition and stoichiometry and determination of molecular sizes of holoenzyme and individual subunits. Later work on Escherichia coli enzyme utilized mutagenesis and optical probes to reveal the molecular mechanism of ATP hydrolysis and detailed facets of catalysis. The second ATPase is P-glycoprotein, which confers multidrug resistance, notably to anticancer drugs, in mammalian cells. Purification of the protein in large quantity allowed detailed characterization of catalysis, formulation of an alternating sites mechanism, and recently, advances in structural characterization.

Looking for the impact of peer review: does count of funding acknowledgements really predict research impact?

A small number of studies have sought to establish that research papers with more funding acknowledgements achieve higher impact and have claimed that such a link exists because research supported by more funding bodies undergoes more peer review. In this paper, a test of this link is made using recently available data from the Web of Science, a source of bibliographic data that now includes funding acknowledgements. The analysis uses 3,596 papers from a single year, 2009, and a single journal, the Journal of Biological Chemistry. Analysis of this data using OLS regression and two ranks tests reveals the link between count of funding acknowledgements and high impact papers to be statistically significant, but weak. It is concluded that count of funding acknowledgements should not be considered a reliable indicator of research impact at this level. Relatedly, indicators based on assumptions that may hold true at one level of analysis may not be appropriate at other levels.