Biredox ionic liquids: new opportunities toward high performance supercapacitors.

Nowadays commercial supercapacitors are based on purely capacitive storage at the porous carbons that are used for the electrodes. However, the limits that capacitive storage imposes on energy density calls to investigate new materials to improve the capacitance of the device. This new type of electrodes (e.g., RuO2, MnO2…) involves pseudo-capacitive faradaic redox processes with the solid material. Ion exchange with solid materials is, however, much slower than the adsorption process in capacitive storage and inevitably leads to significant loss of power. Faradaic process in the liquid state, in contrast can be similarly fast as capacitive processes due to the fast ion transport. Designing new devices with liquid like dynamics and improved specific capacitance is challenging. We present a new approach to increase the specific capacitance using biredox ionic liquids, where redox moieties are tethered to the electrolyte ions, allowing high redox concentrations and significant pseudo-capacitive storage in the liquid state. Anions and cations are functionalized with anthraquinone (AQ) and 2,2,6,6-tetramethylpiperidinyl-1-oxyl (TEMPO) moieties, respectively. Glassy carbon, carbon-onion, and commercial activated carbon electrodes that exhibit different double layer structures and thus different diffusion dynamics were used to simultaneously study the electrochemical response of biredox ionic liquids at the positive and negative electrode.

Rheumatism and chronic fatigue, the two facets of post-chikungunya disease: the TELECHIK cohort study on Reunion island.

Prolonged fatigue is increasingly reported among chikungunya virus (CHIKV)-infected populations. We investigated the relationships between CHIKV exposure, long-lasting rheumatic musculoskeletal pain (LRMSP) and chronic fatigue. 1094 participants (512 CHIKV seropositive and 582 seronegative) of the TELECHIK population-based cohort were analysed considering the duration of the manifestations throughout an average 2-year follow-up. Weighted prevalence rates and prevalence ratios for LRMSP, idiopathic chronic fatigue (ICF), and chronic fatigue syndrome (CFS)-like illness, both latter syndromes adapted from Centers for Disease Control (CDC)-1994/Fukuda criteria, were compared. Population attributable fractions (PAF) were estimated to assess the contribution of CHIKV infection to each of the three phenotypes. Among 362 adult subjects who had reported either rheumatic pain or fatigue at the onset of the infection, weighted prevalence rates of LRMSP, ICF and CFS-like illness were respectively of 32.9%, 38.7% and 23.9%, and of 8.7%, 8.5% and 7.4% among initially asymptomatic peers (P < 0.01, respectively). Each of the three outcomes was highly attributable to chikungunya (PAF of 43.2%, 36.2% and 41.0%, respectively). In the sub-cohort of CHIKV-infected subjects, LRMSP, ICF and CFS-like illness, which overlapped in 70%, accounted for 53% of the chronic manifestations. In addition to rheumatic disease, chronic fatigue could be considered in caring for patients with chronic chikungunya disease.

Individual and contextual risk factors for chikungunya virus infection: the SEROCHIK cross-sectional population-based study.

The purpose of the study was to weigh the community burden of chikungunya determinants on Reunion island. Risk factors were investigated within a subset of 2101 adult persons from a population-based cross-sectional serosurvey, using Poisson regression models for dichotomous outcomes. Design-based risk ratios and population attributable fractions (PAF) were generated distinguishing individual and contextual (i.e. that affect individuals collectively) determinants. The disease burden attributable to contextual determinants was twice that of individual determinants (overall PAF value 89.5% vs. 44.1%). In a model regrouping both categories of determinants, the independent risk factors were by decreasing PAF values: an interaction term between the reporting of a chikungunya history in the neighbourhood and individual house (PAF 45.9%), a maximal temperature of the month preceding the infection higher than 28.5 °C (PAF 25.7%), a socio-economically disadvantaged neighbourhood (PAF 19.0%), altitude of dwelling (PAF 13.1%), cumulated rainfalls of the month preceding the infection higher than 65 mm (PAF 12.6%), occupational inactivity (PAF 11.6%), poor knowledge on chikungunya transmission (PAF 7.3%) and obesity/overweight (PAF 5.2%). Taken together, these covariates and their underlying causative factors uncovered 80.8% of chikungunya at population level. Our findings lend support to a major role of contextual risk factors in chikungunya virus outbreaks.

Platinum for hydrogen sensing: surface and grain boundary scattering antagonistic effects in Pt@Au core-shell nanoparticle assemblies prepared using a Langmuir-Blodgett method.

Hydrogen resistive sensors are fabricated through the synthesis of a series of Pt@Au core-shell nanoparticles showing various Pt shell thicknesses. Resulting colloids are assembled as hexagonal close-packed 2D monolayers of various dimension characteristics using a simple Langmuir-Blodgett method. The fabricated sensors show attractive hydrogen sensing performances with reversible responses in extended sensing ranges, a good specificity towards H2, short response and recovery times… Sensing measurements and data analyses allow the demonstration of the associated sensing mechanisms. The dissociative chemisorption of H2 and O2 on the Pt surface through a Langmuir-Hinshelwood mechanism leads to the formation of chemisorbed hydrogen and hydroxyl groups. This surface nature change induces the modification of the scattering of the conduction electrons at both the grain surface and intercontacts, tuned by the extent of hydrogen and hydroxyl group coverages. In assemblies made of particles showing thin Pt shells, the predominance of the surface scattering described by the Fuchs-Sondheimer model accounts for the observed conductive responses as the number of involved grain boundaries is limited. In contrast, in assemblies made of particles with thick Pt shells, the scattering at the grain boundaries described by the Mayadas-Shatzkes model mostly contributes to the observed resistive responses. The sensor behavior is balanced by these two antagonistic effects.

HIF-1-driven skeletal muscle adaptations to chronic hypoxia: molecular insights into muscle physiology.

Skeletal muscle is a metabolically active tissue and the major body protein reservoir. Drop in ambient oxygen pressure likely results in a decrease in muscle cells oxygenation, reactive oxygen species (ROS) overproduction and stabilization of the oxygen-sensitive hypoxia-inducible factor (HIF)-1α. However, skeletal muscle seems to be quite resistant to hypoxia compared to other organs, probably because it is accustomed to hypoxic episodes during physical exercise. Few studies have observed HIF-1α accumulation in skeletal muscle during ambient hypoxia probably because of its transient stabilization. Nevertheless, skeletal muscle presents adaptations to hypoxia that fit with HIF-1 activation, although the exact contribution of HIF-2, I kappa B kinase and activating transcription factors, all potentially activated by hypoxia, needs to be determined. Metabolic alterations result in the inhibition of fatty acid oxidation, while activation of anaerobic glycolysis is less evident. Hypoxia causes mitochondrial remodeling and enhanced mitophagy that ultimately lead to a decrease in ROS production, and this acclimatization in turn contributes to HIF-1α destabilization. Likewise, hypoxia has structural consequences with muscle fiber atrophy due to mTOR-dependent inhibition of protein synthesis and transient activation of proteolysis. The decrease in muscle fiber area improves oxygen diffusion into muscle cells, while inhibition of protein synthesis, an ATP-consuming process, and reduction in muscle mass decreases energy demand. Amino acids released from muscle cells may also have protective and metabolic effects. Collectively, these results demonstrate that skeletal muscle copes with the energetic challenge imposed by O2 rarefaction via metabolic optimization.

First screening of Aedes albopictus immunogenic salivary proteins.

Study of the human antibody (Ab) response to Aedes salivary proteins can provide new biomarkers to evaluate human exposure to vector bites. The identification of genus- and/or species-specific proteins is necessary to improve the accuracy of biomarkers. We analysed Aedes albopictus immunogenic salivary proteins by 2D immunoproteomic technology and compared the profiles according to human individual exposure to Ae. albopictus or Ae. aegypti bites. Strong antigenicity to Ae. albopictus salivary proteins was detected in all individuals whatever the nature of Aedes exposure. Amongst these antigenic proteins, 68% are involved in blood feeding, including D7 protein family, adenosine deaminase, serpin and apyrase. This study provides an insight into the repertoire of Ae. albopictus immunogenic salivary proteins for the first time.

[Perceived morbidity and community burden of chikungunya in La Reunion]. / Morbidité perçue et impact communautaire du chikungunya a l'île de La Réunion: enquête de cohorte en population TELECHIK.

BACKGROUND: Persistence of clinical manifestations, especially polyarthralgia and fatigue, is a characteristic feature of chikungunya virus (CHIK-v) infection. The purpose of this study was to measure the impact of prolonged or late-onset manifestations of CHIK-v infection on the self-perceived health of people on Reunion Island. METHODS: This retrospective cohort survey, dubbed TELECHIK survey, was conducted eighteen months after the end of the chikungunya outbreak on a representative random sample from the SEROCHIK population-based survey conducted on Reunion Island. A total of 1094 subjects whose CHIK-v specific IgG antibody status had been documented were interviewed about current symptoms. RESULTS: Analysis of data showed 45% of CHIK+ vs 14% of CHIK- subjects reporting musculoskeletal pain (P < 0.001), 56% vs. 44% reporting fatigue (P = 0.003), 77% vs. 53% reporting cerebral manifestations (P < 0.001), 51% vs. 34% reporting sensorineural impairments (P < 0.001), 18% vs. 13% reporting digestive complaints (P = 0.06), and 38% vs. 32% reporting skin involvement (P = 0.13). The mean delay between infection and interview was two years (range, 15-34 months). Analysis of data after correction for age, gender, body mass index and comorbidity indicated that rheumatic pain, fatigue, cerebral manifestations and sensorineural impairments were more likely in CHIK+ than CHIK- subjects but the likelihood of digestive and skin manifestations was the same. CONCLUSION: With a mean delay of two years after infection, 45% to 77% of CHIK+ subjects reported prolonged or late-onset symptoms attributable to CHIK-v. These results indicate that persistent manifestations of chikungunya infection have a heavy impact on rheumatologic, neurological and sensorineural health.

An interdisciplinary approach to controlling chikungunya outbreaks on French islands in the south-west Indian ocean.

The outbreak of chikungunya that occurred on French Island territories in the southwest Indian Ocean in 2005 and 2006 caused severe morbidity and mortality. In the aftermath, French authorities set up a scientific task force including experts in epidemiology, public health, entomology, virology, immunology, sociology, animal health, community and hospital medicine. The mission of the task force was to conceive and propose research programs needed to increase understanding of the disease and epidemic and to help public health officials in improving epidemic response measures. The purpose of this article is to describe the findings of the task force at the end of its two-year existence and initial outcomes in the the areas studied. Discussion emphasizes topics requiring further study.

Fast and robust hydrogen sensors based on discontinuous palladium films on polyimide, fabricated on a wafer scale.

Fast hydrogen sensors based on discontinuous palladium (Pd) films on supporting polyimide layers, fabricated by a cost-efficient and full-wafer compatible process, are presented. The films, deposited by electron-beam evaporation with a nominal thickness of 1.5 nm, consist of isolated Pd islands that are separated by nanoscopic gaps. On hydrogenation, the volume expansion of Pd brings initially separated islands into contact which leads to the creation of new electrical pathways through the film. The supporting polyimide layer provides both sufficient elasticity for the Pd nanoclusters to expand on hydrogenation and a sufficiently high surface energy for good adhesion of both film and contacting electrodes. The novel order of the fabrication processes involves a dicing step prior to the Pd deposition and stencil lithography for the patterning of microelectrodes. This allows us to preserve the as-deposited film properties. The devices work at room temperature, show response times of a few seconds and have a low power consumption of some tens of nW.

Hydrogen sensors and switches from electrodeposited palladium mesowire arrays.

Hydrogen sensors and hydrogen-activated switches were fabricated from arrays of mesoscopic palladium wires. These palladium "mesowire" arrays were prepared by electrodeposition onto graphite surfaces and were transferred onto a cyanoacrylate film. Exposure to hydrogen gas caused a rapid (less than 75 milliseconds) reversible decrease in the resistance of the array that correlated with the hydrogen concentration over a range from 2 to 10%. The sensor response appears to involve the closing of nanoscopic gaps or "break junctions" in wires caused by the dilation of palladium grains undergoing hydrogen absorption. Wire arrays in which all wires possessed nanoscopic gaps reverted to open circuits in the absence of hydrogen gas.

Effect of acute hypoxia on central fatigue during repeated isometric leg contractions.

To determine whether hypoxia has a direct influence on the central command independently of the working muscles, 16 subjects performed intermittent isometric unilateral knee extensions until exhaustion either in normobaric hypoxia (inspired O(2) fraction=0.11, arterial oxygen saturation approximately 84%) or in normoxia while the knee extensor muscles were exposed to circulatory occlusion with a 250 mmHg cuff. Among the subjects, 11 also performed the tests in hypoxia and normoxia without occlusion. Single electrical stimulations were regularly delivered to the femoral nerve to measure the changes in the knee extensor peak twitch force. With the cuff, the average slope of decrease in peak twitch did not depend on the inspired oxygen fraction. Performance was slightly but significantly lower during hypoxia than in normoxia (8.2+/-2.6 vs 9.4+/-3.1 repetitions, P<0.05) with the cuff on. The number of repetitions was much higher during hypoxia with maintaining leg blood flow (15.6+/-4.5 repetitions) than with circulatory occlusion in normoxia. In conclusion, this study showed that a direct effect of hypoxia in reducing the motor drive to the working muscles exists but this effect is moderate.

Transport Properties of Li-TFSI Water-in-Salt Electrolytes.

Water-in-salts are a new family of electrolytes that may allow the development of aqueous Li-ion batteries. They have a structure that is reminiscent of ionic liquids, and they are characterized by a high concentration of ionic species. In this work, we study their transport properties and how they evolve with concentration by using molecular dynamic simulations. We first focus on the choice of the force field. By comparing the simulated viscosities and self-diffusion coefficients with experimental measurements, we select a set of parameters that reproduces well the transport properties. We then use the selected force field to study in detail the variations of the self and collective diffusivities of all the species as well as the transport number of the lithium ion. We show that correlations between ions and water play an important role over the whole concentration range. In the water-in-salt regime, the anions form a percolating network that reduces the cation-anion correlations and leads to rather large values for the transport number compared to other standard electrolytes.

[Early onset neonatal infections in the South of the Reunion Island: Incidence and use of 2002 ANAES risk criteria]. / Infection bactérienne néonatale précoce dans le sud de la Réunion: incidence et application des critères de risque Anaes 2002.

OBJECTIVES: The aim of this study was to describe the incidence of early onset neonatal infections (EONI) in the southern part of the Reunion Island, and to study the application of ANAES criteria. PATIENTS AND METHODS: A cross-sectional study was made of data collected for all live births having occurred between 1st January 2001 and 31st December 2004. RESULTS: Four hundred and thirty-seven in 16,071 neonates (out of 21,231 live births) presented with a certain or probable EONI, accounting for a regional rate of 20 per thousand (CI95 % 18-23 per thousand). Among 437 EONIs, group B streptococcus (GBS) was reported in 70.5% of the cases (n=308), Gram negative bacteria in 19.9% (n=87), of which nearly two thirds of Escherichia coli (n=56). Applying ANAES criteria led to identify 380 EONIs among 437 proven infections (sensitivity: 87%, specificity: 26%). A logistic regression analysis identified eight EONI predictors for the 7015 neonates for whom the mother GBS screening was documented: GBS positive vaginal culture (OR 4.2; CI95% 3.3-5.4), unexplained preterm birth less than 35 weeks (OR 5.7; CI95% 3.7-8.7), prolonged rupture of membranes greater than or equal to 18 hours (OR 2.1; CI95% 1.4-3.0), maternal fever greater than or equal to 37.8 degrees C (OR 3.2; CI95% 2.3-4.5), fetal tachycardia greater than or equal to 160 ppm (OR 2.7; CI95% 1.8-4.0), and thin (OR 1.6; CI95% 1.2-2.1) or thick meconium-stained amniotic fluid (OR 3.0; CI95% 2.1-4.5) or fetid fluid (OR 14.8; CI95% 4.2-51.8). CONCLUSION: The incidence of EONIS far exceeded that observed in metropolitan France, and the ANAES criteria lack sensitivity and specificity.

Trends and sociocultural factors for childhood overweight in La Reunion Island.

OBJECTIVE: To examine time trends (study 1) and sociocultural factors associated with childhood overweight (study 2) in La Reunion Island. DESIGN: Study 1: a longitudinal analysis of anthropometric data available from health services in the town of Saint-Pierre. Study 2: a case - control study. SUBJECTS: Study 1: 1753 children born between 1977 and 1996. Study 2: 101 six-year old overweight children sex-matched with 101 non-overweight children. MEASUREMENTS: Study 1: overweight at birth (birth weight>4 kg) and at age 4 and 6 using French references and references from the International Obesity Task Force. Study 2: parental and perinatal data, child's lifestyle and representation of food, all collected from the mother. RESULTS: Study 1: a dramatic increase in the prevalence rate of overweight at 4 and at 6 is observed, more severe in girls. Study 2: multivariate logistic regressions showed that mother's overweight was the only variable significantly associated with overweight in both the sexes. The other associated factors were related to sociocultural and family features and sedentarity. Logistic probability functions derived from these data are proposed as a tool for detection of at risk families. CONCLUSION: Our results show the need for a targeted prevention of overweight. We offer a proposal based on education and behaviour modification in La Reunion Island.

High prevalence of chronic kidney disease in La Réunion island and its association with the metabolic syndrome in the non-diabetic population: La Réunion Diabetes (REDIA) Study.

AIM: To estimate the prevalence of chronic kidney disease (CKD) in La Réunion island and to investigate the link with the metabolic syndrome in the non-diabetic population. METHODS: The Réunion Diabetes (REDIA) Study included a random sample of 3600 adults aged 30-69 years. Clinical proteinuria (>200 mg/g creatinine), albuminuria (>or=30 mg/g) and estimated glomerular filtration rate (eGFR) were studied in 920 subjects, 411 of whom had diabetes and 509 who did not. Their relations with the metabolic syndrome (as defined by the US National Cholesterol Education Program Adult Treatment Panel III guidelines) were analyzed among those without diabetes. RESULTS: Age-, gender- and diabetes-standardized prevalence of CKD stage 1 or 2 (proteinuria or albuminuria with eGFR>or=60 mL/min/1.73 m2) was 13.8% and, for CKD stage 3 or more (eGFR<60 ml/min/1.73 m2), 10.7%. The adjusted odds ratios (OR) for proteinuria increased with the number of metabolic syndrome traits: 1.5 (95% confidence interval, 0.4-5.2) in non-diabetic participants with one trait compared with those with no trait, 2.0 (CI 0.6-6.6) for two traits and 4.1 (CI 1.3-12.8) for three or more; corresponding ORs for eGFR<60 ml/min/1.73 m2 were 1.9 (CI 0.8-4.5), 0.9 (CI 0.4-2.4) and 2.2 (CI 0.9-5.1), respectively. Clustering of either high blood pressure and triglyceride levels, or high triglycerides and plasma glucose, or all three, conferred the strongest associations with both clinical proteinuria and low eGFR. CONCLUSIONS: CKD prevalence is high in La Réunion island population, and the metabolic syndrome may help to target early diagnosis of CKD in non-diabetic individuals.

Health perceptions of diabetic patients in the REDIA study.

AIM: To explore the beliefs and perceptions of type 2 diabetic patients in La Réunion where the disease is highly prevalent (17.5% among 30-69 yr old subjects) with a strong link to the metabolic syndrome and nutritional habits. METHODS: Two sets of data were analysed. An 80-item questionnaire explored the perceptions of causal factors, knowledge of complications and therapeutic issues in 331 known diabetic patients included in the REDIA study. The data were completed by semi-structured interviews of 40 diabetic patients in a hospital setting. RESULTS: Perceived causal factors of diabetes are mainly sugar excess, heredity and stress or life events. Weight excess and lack of physical activity are virtually never mentioned as causes. Diabetes is predominantly perceived as potentially acute, with risk of coma and death. Its chronic and progressive nature is not appreciated, and chronic complications are not well understood, especially in poorly educated people. Only 33% of males and 42% of females are willing to change their nutritional habits and the role of fats is largely underestimated (30.2%) although 90% consider physical activity as an effective course of action. Most patients are tardy in the way that they adopt medical recommendations and treatment in the course of the disease. CONCLUSION: These results highlight the discrepancy between medical knowledge and patients' perceptions, especially concerning etiological issues and complications. Lifestyle and therapeutic recommendations are not well understood. Educational activities need to consider the knowledge issues and understanding by patients early in the course of chronic diseases like diabetes.

Crystal structure of the Escherichia coli peptide methionine sulphoxide reductase at 1.9 A resolution.

BACKGROUND: Peptide methionine sulphoxide reductases catalyze the reduction of oxidized methionine residues in proteins. They are implicated in the defense of organisms against oxidative stress and in the regulation of processes involving peptide methionine oxidation/reduction. These enzymes are found in numerous organisms, from bacteria to mammals and plants. Their primary structure shows no significant similarity to any other known protein. RESULTS: The X-ray structure of the peptide methionine sulphoxide reductase from Escherichia coli was determined at 3 A resolution by the multiple wavelength anomalous dispersion method for the selenomethionine-substituted enzyme, and it was refined to 1.9 A resolution for the native enzyme. The 23 kDa protein is folded into an alpha/beta roll and contains a large proportion of coils. Among the three cysteine residues involved in the catalytic mechanism, Cys-51 is positioned at the N terminus of an alpha helix, in a solvent-exposed area composed of highly conserved amino acids. The two others, Cys-198 and Cys-206, are located in the C-terminal coil. CONCLUSIONS: Sequence alignments show that the overall fold of the peptide methionine sulphoxide reductase from E. coli is likely to be conserved in many species. The characteristics observed in the Cys-51 environment are in agreement with the expected accessibility of the active site of an enzyme that reduces methionine sulphoxides in various proteins. Cys-51 could be activated by the influence of an alpha helix dipole. The involvement of the two other cysteine residues in the catalytic mechanism requires a movement of the C-terminal coil. Several conserved amino acids and water molecules are discussed as potential participants in the reaction.

A 'specificity' pocket inferred from the crystal structures of the complexes of aldose reductase with the pharmaceutically important inhibitors tolrestat and sorbinil.

BACKGROUND: Aldose reductase (AR) is an NADPH-dependent enzyme implicated in long-term diabetic complications. Buried at the bottom of a deep hydrophobic cleft, the NADPH coenzyme is surrounded by the conserved hydrophilic residues of the AR active site. The existence of an anionic binding site near the NADP+ has been determined from the structures of the complexes of AR with citrate, cacodylate and glucose-6-phosphate. The inhibitor zopolrestat binds to this anionic site, and in the hydrophobic cleft, after a change of conformation which opens a 'specificity' pocket. RESULTS: The crystal structures of the porcine AR holoenzyme and its complexes with the inhibitors tolrestat and sorbinil have been solved; these structures are important as tolrestat and sorbinil are, pharmaceutically, the most well-studied AR inhibitors. The active site of the holoenzyme was analyzed, and binding of the inhibitors was found to involve two contact zones in the active site: first, a recognition region for hydrogen-bond acceptors near the coenzyme, with three centers, including the anionic site; and second, a hydrophobic contact zone in the active-site cleft, which in the case of tolrestat includes the specificity pocket. The conformational change leading to the opening of the specificity pocket upon tolrestat binding is different to the one seen upon zopolrestat binding; this pocket binds inhibitors that are more effective against AR than against aldehyde reductase. CONCLUSIONS: The active site of AR adapts itself to bind tightly to different inhibitors; this happens both upon binding to the inhibitor's hydrophilic heads, and at the hydrophobic and specificity pockets of AR, which can change their shape through different conformational changes of the same residues. This flexibility could explain the large variety of possible substrates of AR.

Protective effect of myostatin gene deletion on aging-related muscle metabolic decline.

While myostatin gene deletion is a promising therapy to fight muscle loss during aging, this approach induces also skeletal muscle metabolic changes such as mitochondrial deficits, redox alteration and increased fatigability. In the present study, we evaluated the effects of aging on these features in aged wild-type (WT) and mstn knockout (KO) mice. Moreover, to determine whether an enriched-antioxidant diet may be useful to prevent age-related disorders, we orally administered to the two genotypes a melon concentrate rich in superoxide dismutase for 12 weeks. We reported that mitochondrial functional abnormalities persisted (decreased state 3 and 4 of respiration; p<0.05) in skeletal muscle from aged KO mice; however, differences with WT mice were attenuated at old age in line with reduced difference on running endurance between the two genotypes. Interestingly, we showed an increase in glutathione levels, associated with lower lipid peroxidation levels in KO muscle. Enriched antioxidant diet reduced the aging-related negative effects on maximal aerobic velocity and running limit time (p<0.05) in both groups, with systemic adaptations on body weight. The redox status and the hypertrophic phenotype appeared to be beneficial to KO mice, mitigating the effect of aging on the skeletal muscle metabolic remodeling.

Structural and biochemical investigations of the catalytic mechanism of an NADP-dependent aldehyde dehydrogenase from Streptococcus mutans.

The NADP-dependent non-phosphorylating glyceraldehyde-3-phosphate dehydrogenase from Streptococcus mutans (abbreviated Sm-ALDH) belongs to the aldehyde dehydrogenase (ALDH) family. Its catalytic mechanism proceeds via two steps, acylation and deacylation. Its high catalytic efficiency at neutral pH implies prerequisites relative to the chemical mechanism. First, the catalytic Cys284 should be accessible and in a thiolate form at physiological pH to attack efficiently the aldehydic group of the glyceraldehyde-3-phosphate (G3P). Second, the hydride transfer from the hemithioacetal intermediate toward the nicotinamide ring of NADP should be efficient. Third, the nucleophilic character of the water molecule involved in the deacylation should be strongly increased. Moreover, the different complexes formed during the catalytic process should be stabilised. The crystal structures presented here (an apoenzyme named Apo2 with two sulphate ions bound to the catalytic site, the C284S mutant holoenzyme and the ternary complex composed of the C284S holoenzyme and G3P) together with biochemical results and previously published apo and holo crystal structures (named Apo1 and Holo1, respectively) contribute to the understanding of the ALDH catalytic mechanism. Comparison of Apo1 and Holo1 crystal structures shows a Cys284 side-chain rotation of 110 degrees, upon cofactor binding, which is probably responsible for its pK(a) decrease. In the Apo2 structure, an oxygen atom of a sulphate anion interacts by hydrogen bonds with the NH2 group of a conserved asparagine residue (Asn154 in Sm-ALDH) and the Cys284 NH group. In the ternary complex, the oxygen atom of the aldehydic carbonyl group of the substrate interacts with the Ser284 NH group and the Asn154 NH2 group. A substrate isotope effect on acylation is observed for both the wild-type and the N154A and N154T mutants. The rate of the acylation step strongly decreases for the mutants and becomes limiting. All these results suggest the involvement of Asn154 in an oxyanion hole in order to stabilise the tetrahedral intermediate and likely the other intermediates of the reaction. In the ternary complex, the cofactor conformation is shifted in comparison with its conformation in the C284S holoenzyme structure, likely resulting from its peculiar binding mode to the Rossmann fold (i.e. non-perpendicular to the plane of the beta-sheet). This change is likely favoured by a characteristic loop of the Rossmann fold, longer in ALDHs than in other dehydrogenases, whose orientation could be constrained by a conserved proline residue. In the ternary and C284S holenzyme structures, as well as in the Apo2 structure, the Glu250 side-chain is situated less than 4 A from Cys284 or Ser284 instead of 7 A in the crystal structure of the wild-type holoenzyme. It is now positioned in a hydrophobic environment. This supports the pK(a) assignment of 7.6 to Glu250 as recently proposed from enzymatic studies.