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BACKGROUND/OBJECTIVES: Temporalis muscle lengthening myoplasty improves tightening of the lips and rehabilitates smile for patients with congenital facial palsies. Because Moebius syndrome is heterogeneous, a careful evaluation is mandatory before deciding to perform myoplasty. This series shows the role of electromyography for investigating temporalis muscle and trigeminal nerve motor functions. METHODS: We conducted a retrospective study of 18 patients with no upward movements of the labial commissure and absent or unsightly smile. Electromyography was used to study the temporalis muscle bilaterally. Analysis focused on the recruitment pattern of voluntary contraction and electrical silence or activity at rest. Traces were classified as normal, neurogenic, or low-amplitude. Functional outcomes of myoplasty were evaluated by measuring the upward movement of the commissure (mm), and qualified as high (≥10), medium (>5), or little (≤5). RESULTS: Surgery was cancelled for 5 patients with abnormal electromyographic signs, neurogenic (2) or low-amplitude (3). Myoplasty was performed in 7 patients (age: 8-17 years), unilaterally (3) or bilaterally (4). Preoperative electromyogram was normal (3), or showed moderate neurogenic (2) or low-amplitude (2) changes. Follow-up period after surgery was from 2 to 12 years; functional outcomes were high (5), medium (1), or little (1). CONCLUSION: Electromyographic study of the temporalis can detect muscle denervation or atrophy, or dyspraxia, and guide decision to encourage or discourage performing myoplasty, or enhance rehabilitation programme and make the patient aware of possibly modest outcome.
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Electromiografía , Síndrome de Mobius/cirugía , Músculo Temporal/fisiopatología , Adolescente , Niño , Femenino , Humanos , Masculino , Síndrome de Mobius/fisiopatología , Contracción Muscular/fisiología , Reclutamiento Neurofisiológico/fisiología , Estudios Retrospectivos , Sonrisa/fisiología , Nervio Trigémino/fisiopatologíaRESUMEN
OBJECTIVE: The monitoring in the post-anaesthesia care unit (PACU) improves the safety, the comfort and the analgesia of patients. At present, studies suggest the possibility to bypass the PACU according to the principle of fast-tracking (FT). The aim of this study was to evaluate the feasibility and the safety of a simulated protocol of FT after a regional anaesthesia. PATIENTS AND METHODS: Seven hundred patients were prospectively included in this study over a period of 6 months. METHODS: The Withes' scoring system was used for determining when patients could be safely discharged from PACU. We added a variable concerning the monitoring of surgical site. A minimum score of 14 was required on arrival to the PACU to consider a FT. The success rate of blocks, the use of sedation or general anaesthesia were noted. Adverse events were recorded. RESULTS: The success rate of blocks was 93 %. The score was higher than 14 in 98 % of case on arrival to the PACU. Thirteen adverse events were reported before surgery and/or operating room. No adverse events were reported during the stay in the PACU. CONCLUSION: Regional anaesthesia seems to be an appropriate principle to fast-track the PACU. It could be a way to reduce health care costs, and can offer solution for the PACU congestion problem. In France, the fast-tracking is a marginal concept without any support regulatory. An evolution to such a practice could be considered.
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Servicio de Anestesia en Hospital/organización & administración , Anestesia de Conducción , Anestesia de Conducción/normas , Estudios de Factibilidad , Femenino , Humanos , Masculino , Persona de Mediana Edad , Estudios ProspectivosRESUMEN
AIM: Hydroxychloroquine therapy during pregnancy is thought to be safe for foetuses. Normal visual function has been showed on clinical grounds in infants exposed in utero to hydroxychloroquine, but there are few visual neurophysiological data. Our study was designed to assess retina and visual pathways using electroretinogram and visual evoked potentials in a series of infants born to mothers treated by hydroxychloroquine for connective tissue diseases. METHODS: Twenty-one infants (3-7 months of age) were consecutively examined between June 2002 and May 2007. Full-field electroretinogram was recorded by contact lens electrodes and visual evoked potentials were recorded by occipital surface electrodes using flash stimulation in mesopic condition. Analysis was focused on the amplitudes and latencies of the a- and b-waves of electroretinogram and the latency of the P(100) component of visual evoked potentials. RESULTS: Electroretinogram abnormalities were detected in six infants, associated with delayed visual evoked potentials in four of them. CONCLUSION: Early electroretinogram and visual evoked potentials testing evidenced neurophysiological visual disturbances in a subset of infants born to mothers treated by hydroxychloroquine. Systematic clinical and neurophysiological vision testing during childhood is needed to detect possible consequences of antenatal exposure to hydroxychloroquine.
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Antirreumáticos/efectos adversos , Hidroxicloroquina/efectos adversos , Enfermedades del Prematuro/inducido químicamente , Efectos Tardíos de la Exposición Prenatal , Trastornos de la Visión/inducido químicamente , Enfermedades del Tejido Conjuntivo/tratamiento farmacológico , Electrorretinografía , Potenciales Evocados Visuales , Femenino , Humanos , Lactante , Recién Nacido , Recien Nacido Prematuro , Enfermedades del Prematuro/diagnóstico , Embarazo , Retina/efectos de los fármacos , Retina/fisiopatología , Trastornos de la Visión/diagnóstico , Vías Visuales/efectos de los fármacos , Vías Visuales/fisiopatologíaRESUMEN
Organophosphorus chemical warfare agents (nerve agents) are to be feared in military operations as well as in terrorist attacks. Among them, VX (O-ethyl-S-[2-(diisopropylamino)ethyl] methylphosphonothioate) is a low volatility liquid that represents a percutaneous as well as an inhalation hazard if aerosolized. It is a potent irreversible cholinesterase (ChE) inhibitor that causes severe signs and symptoms, including respiratory dysfunction that stems from different mechanisms. VX-induced pulmonary oedema was previously reported in dogs but mechanisms involved are not well understood, and its clinical significance remains to be assessed. An experimental model was thus developed to study VX-induced cardiovascular changes and pulmonary oedema in isoflurane-anaesthetized swine. In the course of this study, we observed a fast and unexpected rebound of plasma ChE activity following inhibition provoked by the intravenous injection of 6 and 12 microg kg(-1) of VX. In whole blood ChE activity, the rebound could stay unnoticed. Further investigations showed that the rebound of plasma esterase activity was neither related to spontaneous reactivation of ChE nor to VX-induced increase in paraoxonase/carboxylesterase activities. A bias in Ellman assay, haemoconcentration or severe liver cytolysis were also ruled out. All in all, these results suggest that the rebound was likely due to the release of butyrylcholinesterase into the blood stream from ChE producing organs. Nature of the organ(s) and mechanisms involved in enzyme release will need further investigations as it may represent a mechanism of defence, i.e. VX scavenging, that could advantageously be exploited.
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Sustancias para la Guerra Química/toxicidad , Inhibidores de la Colinesterasa/toxicidad , Colinesterasas/sangre , Compuestos Organotiofosforados/toxicidad , Animales , Butirilcolinesterasa/sangre , Relación Dosis-Respuesta a Droga , Inyecciones Intravenosas , Masculino , Edema Pulmonar/inducido químicamente , Edema Pulmonar/metabolismo , Edema Pulmonar/patología , PorcinosRESUMEN
While there is a consensus that human PON1 (paraoxonase-1) has a protective role, its primary biological function remains unclear. A protective role against poisoning by organophosphates [OPs (organophosphorus compounds)] drove earlier works. Clinical interest has recently focused on a protective role of PON1 against vascular diseases. PON1 resides mainly on HDL (high-density lipoprotein) particles, and converging recent works show that both its activities and stability dramatically depend on this versatile and dynamic molecular environment. The discovery that HPBP (human phosphate-binding protein) has a firm tendency to associate with PON1 has steered new directions for characterizing PON1 functional state(s). Storage stability studies provided evidence that HPBP is involved in maintaining physiologically active PON1 conformation(s). Thermal stability studies showed that human PON1 is remarkably thermostable and that its association with HPBP strongly contributes to slowing down the denaturation rate. A hybrid PON1, displaying mutations that stabilized recombinant enzyme expressed in Escherichia coli, was shown to be more thermostable than natural human PON1. Predictably, its stability was unaffected by the presence of HPBP. Synergistic efforts on characterizing natural PON1 and rPON1 (recombinant PON1) provide information for the design of future stable mutants of PON1-based bioscavengers to be used as safe and effective countermeasures to challenge OPs. Maintaining a stable environment for such administrable human rPON1 should, at least, preserve the anti-atherogenic activity of the enzyme.
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Arildialquilfosfatasa/química , Proteínas de Unión a Fosfato/química , Sitios de Unión , Estabilidad de Enzimas , Humanos , Temperatura , Factores de TiempoRESUMEN
We present particle spectra for charged hadrons pi(+/-), K(+/-), p, and p[over] from pp collisions at square root[s] = 200 GeV measured for the first time at forward rapidities (2.95 and 3.3). The kinematics of these measurements are skewed in a way that probes the small momentum fraction in one of the protons and large fractions in the other. Large proton to pion ratios are observed at values of transverse momentum that extend up to 4 GeV/c, where protons have momenta up to 35 GeV. Next-to-leading order perturbative QCD calculations describe the production of pions and kaons well at these rapidities, but fail to account for the large proton yields and small p[over]/p ratios.
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AIMS OF THE STUDY: Respiratory-related evoked potentials (RREPs) are a method of recording brain activities in response to respiratory stimuli. Although data in childhood are scarce, the absence of the early P1 component of RREPs has been reported in children with a history of life-threatening asthma. This study was focused on the presence, latencies, and amplitudes of the P1, N1, P2, and N2 components of the RREPs in a paediatric series of asthmatic patients. PATIENTS AND METHODS: RREPs were recorded in 21 patients with stable asthma, age range 8-17 years, 11 healthy children, age range 6-16 years, and 24 healthy adults, age range 20-28 years. The signals from left (C3-Cz) and right (C4-Cz) central (rolandic) location were recorded separately, using surface electrodes. Evoked responses to two series of 80 consecutive mid-inspiratory occlusions were averaged. Recordings were analysed manually. RESULTS: All 4 RREPs components were significantly more often absent in asthmatic children than in healthy children and adults (P1, p=0.01; N1, p=0.008; P2, p=0.008, N2, p=0.01). The latencies and amplitudes of the four components were similar in patients and healthy subjects. CONCLUSION: RREPs components were less frequently present in children with asthma than in healthy subjects. This finding should promote the recording of RREPs in other acute and chronic respiratory diseases in children in order to search for possible electroclinical correlations.
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Asma/fisiopatología , Encéfalo/fisiopatología , Potenciales Evocados Somatosensoriales/fisiología , Mecánica Respiratoria/fisiología , Adolescente , Adulto , Envejecimiento/fisiología , Niño , Electroencefalografía , Femenino , Humanos , Masculino , Mecanorreceptores/fisiologíaRESUMEN
We report the serendipitous discovery of a human plasma phosphate binding protein (HPBP). This 38 kDa protein is co-purified with paraoxonase (PON1). The association between HPON1 and HPBP is modulated by phosphate and calcium concentrations. The HPBP X-ray structure solved at 1.9 A resolution is similar to the prokaryotic phosphate solute-binding proteins (SBPs) associated with ATP binding cassette transmembrane transporters, though phosphate-SBPs have never been characterized or predicted from nucleic acid databases in eukaryotes. However, HPBP belongs to the family of ubiquitous eukaryotic proteins named DING, meaning that phosphate-SBPs are also widespread in eukaryotes. The absence of complete genes for eukaryotic phosphate-SBP from databases is intriguing, but the astonishing 90% sequence conservation of genes between evolutionary distant species suggests that the corresponding proteins play an important function. HPBP is the first identified transporter capable of binding phosphate ions in human plasma. Thus it is thought to become a new predictor and a potential therapeutic agent for phosphate-related diseases such as atherosclerosis.
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Apolipoproteínas/química , Secuencia de Aminoácidos , Animales , Arildialquilfosfatasa/química , Cristalografía por Rayos X , Humanos , Modelos Moleculares , Datos de Secuencia Molecular , Fosfatos/sangreRESUMEN
PURPOSE: To assess ocular and clinical manifestations in patients with Möbius syndrome. METHODS: Twenty-seven patients (26 infants and 1 adult) underwent prospective ophthalmic, clinical, neurological, otorhinological, orthopedic and electrophysiological assessment. Twenty-three patients underwent MRI and 20 patients genetic examination with karyotype. RESULTS: Three of 27 patients with cranial nerve palsies did not satisfy the criteria for Möbius syndrome. All 24 patients with Möbius syndrome had facial palsy. Nineteen patients (79.2%) had limited abduction. Eleven patients (45.9%) presented with esotropia, five patients (20.8%) presented with exotropia or hypertropia. Cranial nerve impairment of the Vth, IXth, XIth, and XIIth nerves was noted in 20 patients (83.4%). Other signs were general motor disability in 14 patients (58.2%), orthopedic abnormalities in eight patients (33.3%), and otorhinological abnormalities in six patients (25%). Electromyographic studies of facial muscles revealed neuromuscular changes in all cases. MRI findings showed hypoplasia of facial nerves in two patients (8.3%). Chromosomal abnormalities were not found. One patient presented an inherited inversion of the sixth chromosome. CONCLUSION: The diagnosis of Möbius syndrome may be difficult in some patients with atypical signs of facial diplegia and other cranial nerve palsies. When diagnosing Möbius syndrome, all ophthalmologic and clinical signs must be applied. Möbius syndrome is more than a cranial nerve or nuclear disorder. It is a syndrome of more complex lower brainstem involvement.
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Oftalmopatías/etiología , Síndrome de Mobius/diagnóstico , Adulto , Niño , Diagnóstico Diferencial , Parálisis Facial/etiología , HumanosRESUMEN
OBJECTIVE: To describe the clinical features of a novel variant of autosomal recessive lower motor neuron disease (LMND) with childhood onset and to map the disease-causing gene. METHODS: The authors performed a clinical study in a large consanguineous African family. After linkage exclusion to SMN1 and SOD1 loci, they performed a genome-wide linkage analysis to map the underlying genetic defect. RESULTS: This novel variant of LMND with childhood onset and autosomal recessive mode of inheritance is characterized by a progressive symmetric and generalized involvement of the musculature. Four of the five affected patients had muscle weakness since age 3, strongly worsening during childhood and leading to generalized tetraplegia in adulthood. Genetic analyses using homozygosity mapping strategy assigned this progressive generalized LMND locus to an interval of 3.9 cM (or 1.5 megabases) on chromosome 1p36, between loci D1S508 and D1S2633 (Z(max) = 3.79 at theta = 0.00 at locus D1S253). This region encloses 27 candidate genes. CONCLUSION: Genetic mapping of a novel rare phenotype of lower motor neuron disease opens the way toward the identification of a new gene involved in motor neuron degeneration, located in the 1p36 chromosomal region.
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Cromosomas Humanos Par 1 , Genes Recesivos , Ligamiento Genético , Enfermedad de la Neurona Motora/genética , Adolescente , Adulto , Niño , Mapeo Cromosómico/métodos , Femenino , Humanos , MasculinoRESUMEN
Incoherent elastic neutron scattering experiments on members of the cholinesterase family were carried out to investigate how molecular dynamics is affected by covalent inhibitor binding and by differences in primary and quaternary structure. Tetrameric native and soman-inhibited human butyrylcholinesterase (HuBChE) as well as native dimeric Drosophila melanogaster acetylcholinesterase (DmAChE) hydrated protein powders were examined. Atomic mean-square displacements (MSDs) were found to be identical for native HuBChE and for DmAChE in the whole temperature range examined, leading to the conclusion that differences in activity and substrate specificity are not reflected by a global modification of subnanosecond molecular dynamics. MSDs of native and soman-inhibited HuBChE were identical below the thermal denaturation temperature of the native enzyme, indicating a common mean free-energy surface. Denaturation of the native enzyme is reflected by a relative increase of MSDs consistent with entropic stabilization of the unfolded state. The results suggest that the stabilization of HuBChE phosphorylated by soman is due to an increase in free energy of the unfolded state due to a decrease in entropy.
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Inhibidores de la Colinesterasa/farmacología , Colinesterasas/química , Soman/farmacología , Acetilcolinesterasa/química , Animales , Sitios de Unión , Fenómenos Biofísicos , Biofisica , Butirilcolinesterasa/química , Catálisis , Inhibidores de la Colinesterasa/química , Dicroismo Circular , Dimerización , Drosophila melanogaster , Entropía , Inhibidores Enzimáticos/farmacología , Glicosilación , Humanos , Hidrógeno , Modelos Estadísticos , Neutrones , Distribución Normal , Fosforilación , Conformación Proteica , Desnaturalización Proteica , Pliegue de Proteína , Estructura Cuaternaria de Proteína , Dispersión de Radiación , Soman/química , Especificidad por Sustrato , Temperatura , Termodinámica , Factores de Tiempo , Rayos Ultravioleta , Agua/químicaRESUMEN
Psychological disturbances are frequently encountered in obese patients. The physician specialised in nutrition should take them into account within the therapeutic project and all along the patient's management. This review describe, according to the main obese patients' profiles (typology), the conditions for an optimal adequation between hopes and expectations from both the patient and the physician, as well as the psychiatrist role within the therapeutic project and during the management period.
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Trastornos Mentales/psicología , Trastornos Mentales/terapia , Obesidad/psicología , Humanos , Trastornos Mentales/etiología , Fenómenos Fisiológicos de la Nutrición , Obesidad/complicacionesRESUMEN
Increased muscle extensibility and passivity characterize infantile hypotonia. It may reveal a peripheral neuromuscular disease as well as a disorder of the central nervous system. Electrodiagnostic studies in newborn and young infants are useful to guide the indication of other complementary investigations. Signs of denervation on needle electromyography strongly suggest infantile spinal muscular atrophy. Electrodiagnostic findings can distinguish rare conditions mimicking spinal muscular atrophy that are obstetrical tetraplegia and severe congenital neuropathies. Nerve conduction velocities are severely slowed in hereditary sensorimotor neuropathies and neurodegenerative disorders. Myopathic changes on needle electromyography are associated with congenital muscular dystrophies and structural or metabolic congenital myopathies. In congenital myotonic dystrophy, myotonic discharges can be recorded in the infant as well as in his/her mother. Myopathic changes may also be detected in collagen disorders, in cases of muscular atrophy secondary to hypomotility or malnutrition, and in patients with congenital myasthenic syndrome. Repetitive nerve stimulations are required to characterize myasthenic syndromes. Finally, normal results of electrodiagnostic studies constitute a relevant information that moves diagnostic procedures to search for central nervous system disorders. Benign congenital hypotonia is a quite rare condition that is diagnosed retrospectively, when hypotonia is strictly isolated and recovers completely before 2 years of age.
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Técnicas de Diagnóstico Neurológico , Electromiografía , Hipotonía Muscular/diagnóstico , Humanos , Lactante , Recién Nacido , Hipotonía Muscular/etiología , Hipotonía Muscular/fisiopatología , Enfermedades Musculares/complicaciones , Enfermedades Musculares/fisiopatología , Enfermedades del Sistema Nervioso/complicaciones , Enfermedades del Sistema Nervioso/fisiopatología , Conducción Nerviosa/fisiologíaRESUMEN
Phosphoprotein enriched in astrocytes of 15 kDa (PEA-15) is an abundant phosphoprotein in primary cultures of mouse brain astrocytes. Its capability to interact with members of the apoptotic and mitogen activated protein (MAP) kinase cascades endows PEA-15 with anti-apoptotic and anti-proliferative properties. We analyzed the in vivo cellular sources of PEA-15 in the normal adult mouse brain using a novel polyclonal antibody. Immunohistochemical assays revealed numerous PEA-15-immunoreactive cells throughout the brain of wild-type adult mice while no immunoreactive signal was observed in the brain of PEA-15 -/- mice. Cell morphology and double immunofluorescent staining showed that both astrocytes and neurons could be cellular sources of PEA-15. Closer examination revealed that in a given area only part of the astrocytes expressed the protein. The hippocampus was the most striking example of this heterogeneity, a spatial segregation restricting PEA-15 positive astrocytes to the CA1 and CA3 regions. A PEA-15 immunoreactive signal was also observed in a few cells within the subventricular zone and the rostral migratory stream. In vivo analysis of an eventual PEA-15 regulation in astrocytes was performed using a model of astrogliosis occurring along motor neurons degeneration, the transgenic mouse expressing the mutant G93A human superoxyde-dismutase-1, a model of amyotrophic lateral sclerosis. We observed a marked up-regulation of PEA-15 in reactive astrocytes that had developed throughout the ventral horn of the lumbar spinal cord of the transgenic mice. The heterogeneous cellular expression of the protein and its increased expression in pathological situations, combined with the known properties of PEA-15, suggest that PEA-15 expression is associated with a particular metabolic status of cells challenged with potentially apoptotic and/or proliferative signals.
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Astrocitos/metabolismo , Encéfalo/metabolismo , Neuronas/metabolismo , Fosfoproteínas/biosíntesis , Células 3T3 , Secuencia de Aminoácidos , Animales , Proteínas Reguladoras de la Apoptosis , Astrocitos/citología , Encéfalo/citología , Células Cultivadas , Femenino , Regulación de la Expresión Génica/fisiología , Humanos , Inmunohistoquímica , Péptidos y Proteínas de Señalización Intracelular , Masculino , Ratones , Ratones Transgénicos , Datos de Secuencia Molecular , Neuronas/citología , Fosfoproteínas/inmunologíaRESUMEN
A major result of incoherent elastic neutron-scattering experiments on protein powders is the strong dependence of the intramolecular dynamics on the sample environment. We performed a series of incoherent elastic neutron-scattering experiments on lyophilized human butyrylcholinesterase (HuBChE) powders under different conditions (solvent composition and hydration degree) in the temperature range from 20 to 285 K to elucidate the effect of the environment on the enzyme atomic mean-square displacements. Comparing D(2)O- with H(2)O-hydrated samples, we were able to investigate protein as well as hydration water molecular dynamics. HuBChE lyophilized from three distinct buffers showed completely different atomic mean-square displacements at temperatures above approximately 200 K: a salt-free sample and a sample containing Tris-HCl showed identical small-amplitude motions. A third sample, containing sodium phosphate, displayed highly reduced mean-square displacements at ambient temperature with respect to the other two samples. Below 200 K, all samples displayed similar mean-square displacements. We draw the conclusion that the reduction of intramolecular protein mean-square displacements on an Angstrom-nanosecond scale by the solvent depends not only on the presence of salt ions but also on their type.
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Butirilcolinesterasa/química , Fenómenos Biofísicos , Biofisica , Tampones (Química) , Óxido de Deuterio , Liofilización , Humanos , Iones , Modelos Estadísticos , Neutrones , Protones , Sales (Química)/química , Dispersión de Radiación , Solventes , Temperatura , Agua/químicaRESUMEN
In this paper, the low-resolution structure of a previously unknown protein copurified with human paraoxonase (PON1) is reported. The structure of this protein was very difficult to solve using classical crystallographic methods. Progress was made using a new phasing method based on topological analysis. From the experimental point of view, this method has the advantage of requiring only a simple low-resolution X-ray data set. The program used and the different steps of the data-processing and phasing procedure are described. The results provided an insight into the failure of previous molecular-replacement attempts. The low-resolution shape of the protein which was presented with confidence is compared with and confirmed by the structure at 1.8 A solved subsequently using classical methods. This work shows that this direct-phasing method could be used systematically in difficult cases: it provides low-resolution structural information comparable with that obtainable by electron microscopy.
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Arildialquilfosfatasa/química , Proteínas Sanguíneas/química , Cristalografía por Rayos X/métodos , Arildialquilfosfatasa/sangre , Arildialquilfosfatasa/aislamiento & purificación , Proteínas Sanguíneas/aislamiento & purificación , Cristalización , Análisis de Fourier , Humanos , Modelos Moleculares , Conformación ProteicaAsunto(s)
Electrodiagnóstico/métodos , Electroencefalografía/métodos , Parálisis Facial/congénito , Parálisis Facial/diagnóstico , Factores de Edad , Parpadeo , Diagnóstico Diferencial , Electrodiagnóstico/normas , Electroencefalografía/normas , Parálisis Facial/clasificación , Parálisis Facial/epidemiología , Humanos , Incidencia , Lactante , Recién Nacido , Actividad Motora , Conducción Nerviosa , Tiempo de Reacción , Estudios Retrospectivos , Índice de Severidad de la EnfermedadRESUMEN
The inactivation process of native (N) human butyrylcholinesterase (BuChE) by pressure and/or heat was found to be multi-step. It led to irreversible formation of an active intermediate (I) state and a denatured state. This series-inactivation process was described by expanding the Lumry-Eyring [Lumry, R. and Eyring, H. (1954) J. Phys. Chem. 58, 110-120] model. The intermediate state (I) was found to have a K(m) identical with that of the native state and a turnover rate (k(cat)) twofold higher than that of the native state with butyrylthiocholine as the substrate. The increased catalytic efficiency (k(cat)/K(m)) of I can be explained by a conformational change in the active-site gorge and/or restructuring of the water-molecule network in the active-site pocket, making the catalytic steps faster. However, a pressure/heat-induced covalent modification of native BuChE, affecting the catalytic machinery, cannot be ruled out. The inactivation process of BuChE induced by the combined action of pressure and heat was found to continue after interruption of pressure/temperature treatment. This secondary inactivation process was termed 'remnant inactivation'. We hypothesized that N and I were in equilibrium with populated metastable N' and I' states. The N' and I' states can either return to the active forms, N and I, or develop into inactive forms, N(')(in) and I(')(in). Both active N' and I' intermediate states displayed different rates of remnant inactivation depending on the pressure and temperature pretreatments and on the storage temperature. A first-order deactivation model describing the kinetics of the remnant inactivation of BuChE is proposed.
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Butirilcolinesterasa/metabolismo , Butirilcolinesterasa/química , Dominio Catalítico , Inhibidores de la Colinesterasa , Calor , Humanos , Técnicas In Vitro , Cinética , Modelos Químicos , Presión , Conformación Proteica , Desnaturalización Proteica , TermodinámicaRESUMEN
The effect of high electric field in capillary zone electrophoresis (CZE) was evaluated for the study of the thermally induced unfolding of Bungarus fasciatus acetylcholinesterase. This monomer enzyme is characterised by two interdependent uncommon structural features, the asymmetrical distribution of charged residues and a relatively low thermal denaturation temperature. Both traits were presumed to interfere in the thermal unfolding of this enzyme as investigated by CZE. This paper analyses the effect of high electric field on the behaviour of the enzyme native state. It is shown that increasing the applied field causes denaturation-like transition of the enzyme at a current power which does not induce excessive Joule heating in the capillary. The susceptibility to electric field of proteins like cholinesterases, with charge distribution anisotropy, large permanent dipole moment and notable molecular flexibility associated with moderate thermal stability, was subsequently discussed.