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1.
Metrologia ; 59(4)2022.
Article in English | MEDLINE | ID: mdl-36733421

ABSTRACT

We have developed generalized methods for electrical substitution optical measurements, as well as cryogenic detectors which can be used to implement them. The new methods detailed here enable measurement of arbitrary periodic waveforms by an electrical substitution radiometer (ESR), which means that spectral and dynamic optical power can be absolutely calibrated directly by a primary standard detector. Cryogenic ESRs are not often used directly by researchers for optical calibrations due to their slow response times and cumbersome operation. We describe two types of ESRs with fast response times, including newly developed cryogenic bolometers with carbon nanotube absorbers, which are manufacturable by standard microfabrication techniques. These detectors have response times near 10 ms, spectral coverage from the ultraviolet to far-infrared, and are ideal for use with generalized electrical substitution. In our first tests of the generalized electrical substitution method with FTS, we have achieved uncertainty in detector response of 0.13 % (k=1) and total measurement uncertainty of 1.1 % (k=1) in the mid-infrared for spectral detector responsivity calibrations. The generalized method and fast detectors greatly expand the range of optical power calibrations which can be made using a wideband primary standard detector, which can shorten calibration chains and improve uncertainties.

2.
Opt Express ; 29(23): 37314-37326, 2021 Nov 08.
Article in English | MEDLINE | ID: mdl-34808806

ABSTRACT

We have demonstrated the first continuous-scan electrical substitution Fourier transform spectrometer (ES-FTS), which serves initially as an apparatus for absolute spectral responsivity calibrations of detectors over the wavelength range from 1.5 µm to 11 µm. We present data on the realization of a spectral detector-comparator system with high accuracy, high dynamic range, high spectral resolution and fast measurement in the infrared region, which is tied directly to an absolute power scale through electrical substitution. The ES-FTS apparatus employs a commercial Fourier transform spectrometer and a custom electrical substitution bolometer detector to enable spectrally-resolved absolute optical power measurements. A generalization of electrical substitution techniques enables determination of the voltage waveform that must be applied to the bolometer's electrical heater to cancel the optical signal from a Michelson interferometer in order to quantify the time-dependent optical power incident on the bolometer. The noise floor of the electrical substitution bolometer is on the order of 10 pW/Hz½ and its response is expected to be linear from the noise floor to 1 mW. A direct comparison between a pyroelectric standard detector and the ES-FTS has been performed, and experimental results reported here show great potential for this technique.

3.
Appl Opt ; 57(18): D82-D89, 2018 Jun 20.
Article in English | MEDLINE | ID: mdl-30117943

ABSTRACT

We have designed, fabricated, and measured infrared trap detectors made from arsenic-doped silicon (Si:As) blocked impurity band (BIB) photodetectors. These trap detectors are composed of two detectors in a wedge geometry, with an entrance aperture diameter of either 1 or 3 mm. The detectors were calibrated for quantum efficiency against a pyroelectric reference detector using a Fourier transform spectral comparator system, and etalon effects and spatial uniformity of the traps were also quantified. Measurements of the traps at a temperature of 10 K show that nearly ideal external quantum efficiency (>90%) can be attained over much of the range from 4 to 24 µm, with significant responsivity from 2 to 30 µm. The traps exhibited maximum etalon oscillations of only 2%, which is about 10 times smaller amplitude than those of the single Si:As BIB detectors measured under similar conditions. Spatial nonuniformity across the entrance apertures of the traps was about 1%. The combination of high detectivity, wideband wavelength coverage, spectral flatness, and spatial uniformity make these trap detectors an excellent reference detector for spectrally resolved measurements and radiometric calibrations over the near- to far-infrared wavelength range.

4.
Acta Ortop Mex ; 31(3): 148-151, 2017.
Article in Spanish | MEDLINE | ID: mdl-29216707

ABSTRACT

The concept of arthrogryposis encompasses several conditions that share the presence of multiple congenital contractures. The knee is frequently involved and is an important cause of morbidity in these patients. Flexion contractures of the knee are the most common ones and have a worse prognosis than extension contractures. Different approaches are available to treat flexion contractures of the knee. Distal femoral extension osteotomy effectively corrects fixed flexion, but may lead to residual deformity. This iatrogenic deformity disrupts the anterior convexity of the femoral shaft and leads to serious problems in the subsequent management of orthopedic conditions. This is a case report of a patient with arthrogryposis and a femur deformity who sustained a supracondylar fracture. Managing the fracture was challenging due to a disruption in the normal architecture of the femur. This is a description of the osteosynthesis approach used to treat the fracture and the patients pre-existing deformity.


El concepto «artrogriposis¼ agrupa varias enfermedades cuya característica común es la presencia de múltiples contracturas congénitas. La rodilla se afecta con frecuencia y es causa importante de morbilidad en estos pacientes. Las contracturas en flexión de la rodilla son más comunes y tienen peor pronóstico que las contracturas en extensión. Existen diferentes técnicas de tratamiento para la contractura en flexión de la rodilla; la osteotomía distal de extensión femoral corrige eficazmente la flexión fija, pero puede dar lugar a una deformidad residual. Esta deformidad iatrogénica altera la convexidad anterior de la diáfisis femoral y conlleva problemas graves en el manejo posterior de patología ortopédica. Se presenta el caso de una paciente con artrogriposis y una deformidad en fémur con una fractura en la región supracondílea. El manejo de la fractura fue dificultoso por la presencia de una distorsión de la arquitectura normal del fémur. En nuestro trabajo se describe el método de osteosíntesis utilizado para hacer frente a la fractura y a la deformidad previa de la paciente.


Subject(s)
Arthrogryposis , Femoral Fractures , Fracture Fixation, Internal , Arthrogryposis/complications , Femoral Fractures/etiology , Femoral Fractures/surgery , Femur , Fracture Fixation, Internal/methods , Humans , Knee Joint , Osteotomy , Range of Motion, Articular
5.
Acta ortop. mex ; 31(3): 148-151, may.-jun. 2017. graf
Article in Spanish | LILACS | ID: biblio-886555

ABSTRACT

Resumen: El concepto «artrogriposis¼ agrupa varias enfermedades cuya característica común es la presencia de múltiples contracturas congénitas. La rodilla se afecta con frecuencia y es causa importante de morbilidad en estos pacientes. Las contracturas en flexión de la rodilla son más comunes y tienen peor pronóstico que las contracturas en extensión. Existen diferentes técnicas de tratamiento para la contractura en flexión de la rodilla; la osteotomía distal de extensión femoral corrige eficazmente la flexión fija, pero puede dar lugar a una deformidad residual. Esta deformidad iatrogénica altera la convexidad anterior de la diáfisis femoral y conlleva problemas graves en el manejo posterior de patología ortopédica. Se presenta el caso de una paciente con artrogriposis y una deformidad en fémur con una fractura en la región supracondílea. El manejo de la fractura fue dificultoso por la presencia de una distorsión de la arquitectura normal del fémur. En nuestro trabajo se describe el método de osteosíntesis utilizado para hacer frente a la fractura y a la deformidad previa de la paciente.


Abstract: The concept of arthrogryposis encompasses several conditions that share the presence of multiple congenital contractures. The knee is frequently involved and is an important cause of morbidity in these patients. Flexion contractures of the knee are the most common ones and have a worse prognosis than extension contractures. Different approaches are available to treat flexion contractures of the knee. Distal femoral extension osteotomy effectively corrects fixed flexion, but may lead to residual deformity. This iatrogenic deformity disrupts the anterior convexity of the femoral shaft and leads to serious problems in the subsequent management of orthopedic conditions. This is a case report of a patient with arthrogryposis and a femur deformity who sustained a supracondylar fracture. Managing the fracture was challenging due to a disruption in the normal architecture of the femur. This is a description of the osteosynthesis approach used to treat the fracture and the patient's pre-existing deformity.


Subject(s)
Humans , Arthrogryposis/complications , Femoral Fractures/surgery , Femoral Fractures/etiology , Fracture Fixation, Internal/methods , Osteotomy , Range of Motion, Articular , Femur , Knee Joint
8.
Lancet ; 380(9849): 1229-30, 2012 Oct 06.
Article in English | MEDLINE | ID: mdl-23040858

Subject(s)
Research/economics , Humans
9.
J Chromatogr B Analyt Technol Biomed Life Sci ; 879(29): 3277-82, 2011 Nov 01.
Article in English | MEDLINE | ID: mdl-21515096

ABSTRACT

The availability of enzymes with a high promiscuity/specificity relationship permits the hydrolysis of several substrates with a view to obtaining a certain product or using one enzyme for several productive lines. N-Carbamoyl-ß-alanine amidohydrolase from Agrobacterium tumefaciens (Atßcar) has shown high versatility to hydrolyze different N-carbamoyl-, N-acetyl- and N-formyl-amino acids to produce different α, ß, γ and δ amino acids. We have calculated the promiscuity index for the enzyme, obtaining a value of 0.54, which indicates that it is a modestly promiscuous enzyme. Atßcar presented the highest probability of hydrolysis for N-carbamoyl-amino acids, being the enzyme more efficient for the production of α-amino acids. We have also demonstrated by mutagenesis, modelling, kinetic and binding experiments that W218 and A359 indirectly influence the plasticity of the enzyme due to interaction with the environment of R291, the key residue for catalytic activity.


Subject(s)
Agrobacterium tumefaciens/enzymology , Amidohydrolases/chemistry , Amidohydrolases/metabolism , Amino Acids/biosynthesis , beta-Alanine/metabolism , Agrobacterium tumefaciens/genetics , Chromatography, High Pressure Liquid , Circular Dichroism , Elasticity , Escherichia coli/genetics , Escherichia coli/metabolism , Kinetics , Models, Molecular , Mutagenesis, Site-Directed , Protein Binding , Recombinant Proteins/chemistry , Recombinant Proteins/metabolism , Stereoisomerism , Substrate Specificity , Thermodynamics , beta-Alanine/chemistry
10.
Med. intensiva ; 28(4)2011. ilus, tab
Article in Spanish | LILACS | ID: biblio-908957

ABSTRACT

Objetivo. Evaluar el impacto del uso de un protocolo de medidas de detección y resucitación precoz durante la pandemia de gripe A (H1N1) sobre el ingreso de pacientes en el Servicio de Terapia Intensiva (STI). Diseño. Estudio de observación y retrospectivo de pacientes críticos. Ámbito. Un STI médico-quirúrgico. Pacientes. Adultos que ingresaron en el STI desde el Servicio de Urgencia entre el 28 de abril de 2009 y el 4 de septiembre de 2009. Variables de interés principal. Sexo, edad, comorbilidades, APACHE II, SOFA al ingreso en el STI, sintomatología clínica, imágenes radiográficas, necesidad de asistencia respiratoria mecánica (ARM), ácido láctico (AL), creatinfosfoquinasa (CPK), lactato deshidrogenasa (LDH), leucocitosis, saturación de O2 (SatO2) y PaO2/FiO2 al ingresar en el STI, presión al final de la espiración (PEEP), días de ARM, tratamiento con oseltamivir (dosis/tiempo), aislamiento bacteriológico y virológico en secreción bronquial, tratamiento con corticoides, estadía en el STI y mortalidad. Resultados. Se incluyeron 13 pacientes que ingresaron en el STI durante el período estudiado; media de la edad 45 ± 3; mujeres: 8 (61,5%), comorbilidades (n = 7, 53,8%): enfermedad pulmonar obstructiva crónica (n = 3), diabetes (n = 2), insuficiencia cardíaca (n = 1), cirrosis (n = 1), APACHE II: 18, SOFA: 9 ± 2. La sintomatología clínica predominante fue la siguiente: fiebre (n = 13, 100%), tos (n = 11, 84,6%), disnea (n = 9, 69,2%), infiltrados intersticiales (5/13, 38,4%), opacidades alveolares (6/13, 46,1%), opacidades mixtas (2/13, 15,3%), cuatro cuadrantes (9/13, 69,2%) y dos cuadrantes (4/13, 30,7%); se hallaron los siguientes valores medios: AL 25 mg/dl, CPK 480 U/l (p <0,05), LDH 2100 U/l (p <0,001), leucocitosis 12.500 mm3 , PEEP 18 cm H2O, SatO2 <91% (n = 11, 84,6%), PaO2/FiO2 <150 (n = 11, 84,6%), necesidad de ARM (n = 11, 84,6%), días de ARM 9,5 ± 3 días. Oseltamivir: dosis 150 mg/12 h; aislamiento bacteriológico: neumococo (n = 7, 53,8%); aislamiento virológico: H1N1 (n = 5, 38,4%); duración: 9,5 ± 3 días, corticoides (n = 8, 61,5%). Tiempo en el STI: 11 ± 4. No hubo muertes.    Conclusión. La aplicación de un protocolo inicial en el que además se evalúo la gravedad benefició la correcta evaluación y resucitación inicial en el grupo estudiado. Los valores de CPK y de LDH se acompañaron de hipoxemia severa y mayor compromiso pulmonar en la radiología de tórax(AU)


Objective. To evaluate the impact of a protocol for screening and resuscitation measures during the pandemic H1N1 on patients´ admission to the Intensive Care Unit (ICU). Design. Retrospective observational study of critically ill patients. Place. A medical-surgical Intensive Care Unit (ICU). Patients. Adults admitted to the ICU from the Emergency Department from April 28th 2009 to September 4th 2009. Variables of primary interest. Sex, age, comorbidities, APACHE II, SOFA at admission to the ICU, clinical symptoms, radiographic images, need for mechanical ventilation (MV), lactic acid (AL), creatine kinase (CPK), lactate dehydrogenase (LDH), leukocytosis, O2 saturation (O2Sat) and PaO2/FiO2 at admission to the ICU, end-expiratory pressure (PEEP), days of MV (DMV), oseltamivir (dose/time), bacteriological and virological isolations in bronchial secretions, corticosteroid treatment, stay in the ICU and mortality. Results. We included 13 patients admitted to ICU during the study period; mean age: 45 ± 3; females: 8 (61.5%); comorbidities (n = 7, 53.8%): COPD (n = 3), diabetes (n = 2), heart failure (n = 1), cirrhosis (n = 1), APACHE II: 18, SOFA: 9 ±-2. Clinical symptoms were: fever (n = 13, 100%), cough (n = 11, 84.6%), dyspnea (n = 9, 69.2%), interstitial infiltrates (5/13, 38.4%), alveolar opacities (6/13, 46.1%), mixed opacities (2/13, 15.3%), four quadrants (9/13, 69.2%) and two quadrants (4/13, 30.7%), average measures: AL 25 mg/dL, CPK 480 U/L (p <0.05), LDH 2,100 U/L (p <0.001), leukocytosis 12,500 mm3 , PEEP 18 cm H2O, O2Sat <91% (n = 11, 84.6%), PaO2/FiO2 <150 (n = 11, 84.6%), MV (n = 11, 84.6%), DMV: 9.5 ± 3; oseltamivir: 150 mg/12 hours; bacteriological isolation (Pneumococcus: n = 7, 53.8%), virological isolation (H1N1: n = 5, 38.4%); length: 9.5 ± 3 days, corticosteroids (n = 8, 61.5%); ICU stay 11 ± 4; no deaths were reported. Conclusion. The application of a protocol in which severity was also evaluated benefited in the correct assessment and initial resuscitation. The values of CPK and LDH were associated with severe hypoxemia and lung involvement in the thorax xrays. (AU)


Subject(s)
Humans , Resuscitation , Influenza A Virus, H1N1 Subtype
11.
Theriogenology ; 73(5): 595-604, 2010 Mar 15.
Article in English | MEDLINE | ID: mdl-20035987

ABSTRACT

This study examined the influence of the following growth factors and cytokines on early embryonic development: insulin-like growth factors I and II (IGF-I, IGF-II), basic fibroblast growth factor (bFGF), transforming growth factor (TGF-beta), granulocyte-macrophage colony-stimulating factor (GM-CSF), and leukemia inhibitory factor (LIF). Synthetic oviduct fluid (SOF) was used as the culture medium. We studied the development of bovine embryos produced in vitro and cultured until Day 9 after fertilization. TGF-beta1, bFGF, GM-CSF, and LIF used on their own significantly improved the yield of hatched blastocysts. IGF-I, bFGF, TGF-beta1, GM-CSF, and LIF significantly accelerated embryonic development, especially the change from the expanded blastocyst to hatched blastocyst stages. Use of a combination of these growth factors and cytokines (GF-CYK) in SOF medium produced higher percentages of blastocysts and hatched blastocysts than did use of SOF alone (45% and 22% vs. 24% and 12%; P<0.05) on Day 8 after in vitro fertilization and similar results to use of SOF+10% fetal calf serum (38% and 16%, at the same stages, respectively). The averages of total cells, inner cell mass cells, and trophectoderm cells of exclusively in vitro Day-8 blastocysts for pooled GF-CYK treatments were higher than those for SOF and similar to those for fetal calf serum. The presence of these growth factors and cytokines in the embryo culture medium therefore has a combined stimulatory action on embryonic development; in particular through an increase in hatching rate and in the number of cells of both the inner cell mass and trophoblast. These results are the first to demonstrate that use of a combination of recombinant growth factors and cytokine, as IGF-I, IGF-II, bFGF, TGF-beta1, LIF, and GM-CSF, produces similar results to 10% fetal calf serum for the development of in vitro-produced bovine embryos. This entirely synthetic method of embryo culture has undeniable advantages for the biosecurity of embryo transfer.


Subject(s)
Embryonic Development/drug effects , Fibroblast Growth Factor 2/administration & dosage , Granulocyte-Macrophage Colony-Stimulating Factor/administration & dosage , Insulin-Like Growth Factor II/administration & dosage , Insulin-Like Growth Factor I/administration & dosage , Leukemia Inhibitory Factor/administration & dosage , Transforming Growth Factor beta1/administration & dosage , Animals , Cattle , Culture Media, Serum-Free/pharmacology , Drug Combinations , Embryo Culture Techniques , Embryo, Mammalian , Female , Fertilization in Vitro , Fibroblast Growth Factor 2/pharmacology , Granulocyte-Macrophage Colony-Stimulating Factor/pharmacology , Insulin-Like Growth Factor I/pharmacology , Insulin-Like Growth Factor II/pharmacology , Leukemia Inhibitory Factor/pharmacology , Male , Transforming Growth Factor beta1/pharmacology
12.
Reprod Domest Anim ; 42(1): 68-75, 2007 Feb.
Article in English | MEDLINE | ID: mdl-17214777

ABSTRACT

The interferon-tau (IFN-tau) secretion levels after hatching by bovine blastocysts derived from in vitro maturated oocytes (Group A) and from in vivo (Group B) were investigated considering embryo quality. Only very homogeneous blastocysts of excellent or good quality were considered from day 7 of culture (Group A) and day 7 after artificial insemination with frozen-thawed from the same bull used for in vitro fertilization (Group B). All embryos were individually cultured into a 50 microl droplet of synthetic oviduct fluid medium with 10% fetal calf serum. After 24-h culture both Group A (n=44) and B (n=40) secreted <54 pm IFN-tau. After 48-, 72-, 96- and 120-h culture, Group A daily secreted 143 +/- 24 pm IFN-tau (n=19) vs 85 +/- 12 pm IFN-tau (n=21) for Group B (p < 0.01), 491 +/- 128 pm IFN-tau (n=29) vs 216 +/- 37 pm IFN-tau (n=23) (NS), 499 +/- 135 pm IFN-tau (n=26) vs 353 +/- 93 pm IFN-tau (n=21) (NS), 559 +/- 136 pm IFN-tau (n=22) vs 333 +/- 75 pm IFN-tau (n=20) (NS), respectively. Taken all together during 5 days, Group A produced per embryo 1690 +/- 290 pm IFN-tau (n=22) vs 982 +/- 182 pm IFN-tau (n=20) for Group B (p < 0.05). For all culture time there were sizable percentages of embryos that did not produce concentrations of IFN-tau above a certain cut-off level, and as such were not used to compute the means. In respect of the embryo quality whatever the groups after days 7-12 of culture, IFN-tau secretions were 1815 +/- 453 pm (n=10) for the embryos of excellent quality vs 1356 +/- 200 pm (n=28) for those of good quality (NS) and 360 +/- 188 pm (n=4) (p < 0.05) for embryos of fair quality. A positive relationship between IFN-tau production and in vitro development of quality I embryos was observed, whatever the embryos origins and, the embryos completely produced in vitro secreted more IFN-tau than the embryos produced in vivo.


Subject(s)
Blastocyst/metabolism , Cattle/embryology , Interferon Type I/metabolism , Pregnancy Proteins/metabolism , Animals , Antiviral Agents/metabolism , Cell Count/veterinary , Female , Fertilization in Vitro/veterinary , In Vitro Techniques , Male , Pregnancy , Time Factors
13.
J Res Natl Inst Stand Technol ; 111(5): 401-10, 2006.
Article in English | MEDLINE | ID: mdl-27274942

ABSTRACT

A common radiometric platform for the development of application-specific metrics to quantify the performance of sensors and systems is described. Using this platform, sensor and system performance may be quantified in terms of the accuracy of measurements of standardized sets of source distributions. The prototype platform consists of spectrally programmable light sources that can generate complex spectral distributions in the ultraviolet, visible and short-wave infrared regions for radiometric, photometric and colorimetric applications. In essence, the programmable spectral source is a radiometric platform for advanced instrument characterization and calibration that can also serve as a basis for algorithm testing and instrument comparison.

14.
Protein Pept Lett ; 12(2): 123-37, 2005 Feb.
Article in English | MEDLINE | ID: mdl-15723638

ABSTRACT

The small size and lack of disulphide bonds or cofactors in the Histidine-containing phosphocarrier protein (HPr) makes it an attractive system with which to study structure, interaction to its enzymatic partners, and its stability and folding. Here we give an overview on the immense work that has been performed on this protein and we will show that HPr has been widely used as a model protein to study important aspects in modern Structural Biology.


Subject(s)
Antigens, Bacterial/chemistry , Bacterial Proteins/chemistry , Phosphoenolpyruvate Sugar Phosphotransferase System/chemistry , Protein Folding , Antigens, Bacterial/genetics , Antigens, Bacterial/metabolism , Bacterial Proteins/genetics , Bacterial Proteins/metabolism , Binding Sites , Enzyme Stability , Histidine/metabolism , Hydrogen Bonding , Models, Molecular , Phosphoenolpyruvate Sugar Phosphotransferase System/genetics , Phosphoenolpyruvate Sugar Phosphotransferase System/metabolism , Protein Conformation , Protein Denaturation , Serine/metabolism
15.
Anal Bioanal Chem ; 378(2): 447-55, 2004 Jan.
Article in English | MEDLINE | ID: mdl-14598009

ABSTRACT

Palladium, iridium, and rhodium are evaluated as possible chemical modifiers in the determination of As in digest solutions of biological materials (human hair and clam) by tungsten coil electrothermal atomic absorption spectrophotometry (TCA-AAS). The modifier in solution was applied onto the coil and thermally pre-reduced; the pre-reduction conditions, the amount of modifier, and the thermal program were optimized. Palladium was not satisfactory, whereas Ir and Rh were effective modifiers and rendered better relative sensitivity for As by a factor of 1.4 and 1.9, respectively compared to the case without modifier. Upon optimization of thermal conditions for As in pre-reduced Ir (2.0 microg) and Rh (2.0 microg) modifiers and in the digest solutions of the study matrices, Rh (2.0 microg) was more effective modifier and was selected as such. The mean within-day repeatability was 2.8% in consecutive measurements (25-100 microg L(-1)) (3 cycles, each of n=6) and confirmed good short-term stability of the absorbance measurements. The mean reproducibility was 4.4% (n=20 in a 3-day period) and the detection limit (3 sigmablank/slope) was 29 pg (n=15). The useful coil lifetime in Rh modifier was extended to 300-400 firings. Validation was by determination of As in the certified reference material (CRM) of "Oyster tissue" solution with a percentage relative error (Erel%) of 2% and percentage relative standard deviation (RSD%) of 3% (n=4), and by analytical recovery of As spiked in CRM of human hair [94 +/- 8% (n=4)]. The methodology is simple, fast (sample readout frequency 21 h(-1)), reliable, of low cost, and was applied to the determination of As in hair samples of exposed and unexposed workers.


Subject(s)
Arsenic/analysis , Iridium/chemistry , Palladium/chemistry , Rhodium/chemistry , Spectrophotometry, Atomic/methods , Tungsten , Animals , Bivalvia/chemistry , Hair/chemistry , Humans , Spectrophotometry, Atomic/instrumentation , Temperature
16.
Biochemistry ; 40(50): 15246-56, 2001 Dec 18.
Article in English | MEDLINE | ID: mdl-11735407

ABSTRACT

Proton NMR spectral resonances of thioredoxin m from spinach have been assigned, and its solution structure has been determined on the basis of 1156 nuclear Overhauser effect- (NOE-) derived distance constraints by using restrained molecular dynamics calculations. The average pairwise root-mean-square deviation (RMSD) for the 25 best NMR structures for the backbone was 1.0 +/- 0.1, when the structurally well-defined residues were considered. The N- and C-terminal segments (1-13 and 118-119) and residues 41-49, comprising the active site, are highly disordered. At the time of concluding this work, a crystal structure of this protein was reported, in which thioredoxin m was found to crystallize as noncovalent dimers. Although the solution and crystal structures are very similar, no evidence was found about the existence of dimers in solution, thus confirming that dimerization is not needed for the regulatory activity of thioredoxin m. The spinach thioredoxin m does not unfold by heat in the range 25-85 degrees C, as revealed by thermal circular dichroic (CD) measurements. However, its unfolding free energy (9.1 +/- 0.8 kcal mol(-1), at pH 5.3 and 25 degrees C) could be determined by extrapolating the free energy values obtained at different concentrations of guanidinium chloride (GdmCl). The folding-unfolding process is two-state as indicated by the coincidence of the CD denaturation curves obtained at far and near UV. The H/D exchange behavior of backbone amide protons was analyzed. The slowest-exchanging protons, requiring a global-unfolding mechanism in order to exchange, are those from beta2, beta3, and beta4, the central strands of the beta-sheet, which constitute the main element of the core of the protein. The free energies obtained from exchange measurements of protons belonging to the alpha-helices are lower than those derived from GdmCl denaturation studies, indicating that those protons exchange by local-unfolding mechanisms.


Subject(s)
Thioredoxins/chemistry , Amides/chemistry , Amino Acid Sequence , Circular Dichroism , Crystallization , Crystallography, X-Ray , Dimerization , Drug Stability , Magnetic Resonance Spectroscopy , Models, Molecular , Molecular Sequence Data , Oxidation-Reduction , Protein Folding , Protein Structure, Secondary , Protons , Solutions , Spinacia oleracea/chemistry , Spinacia oleracea/genetics , Thermodynamics , Thioredoxins/genetics
17.
Eur J Biochem ; 268(18): 4868-77, 2001 Sep.
Article in English | MEDLINE | ID: mdl-11559355

ABSTRACT

We have analysed the hydrogen/deuterium exchange of the tetramerization domain of human tumour suppressor p53 under mild chemical denaturation conditions, and at different temperatures. Exchange behaviour has been measured for 16 amide protons in the chemical-denaturation studies and for seven protons in the temperature-denaturation studies. The exchange rates are in the range observed for other proteins with similar elements of secondary structure. The slowest-exchange core includes contributions from residues in the alpha helix and the beta sheet. However, only some of the slowest-exchanging protons correspond to residues involved in native interactions in the transient intermediate detected during the folding of this domain. The guanidinium-chloride denaturation curves of all residues seem to merge together, although they are well below the main isotherm of global unfolding. Thus, there is no evidence for several subglobal unfolding units. The activation parameters obtained from the temperature-denaturation experiments are similar to those obtained for monomeric proteins, and well below the global unfolding enthalpy obtained by circular dichroism measurements. Thus, the exchange studies at different denaturant concentrations and temperatures indicate that no particular folding intermediate is populated under those conditions.


Subject(s)
Hydrogen/metabolism , Tumor Suppressor Protein p53/chemistry , Tumor Suppressor Protein p53/metabolism , Dimerization , Guanidine/pharmacology , Humans , Hydrogen Bonding , Kinetics , Magnetic Resonance Spectroscopy , Models, Molecular , Protein Denaturation/drug effects , Protein Folding , Protein Structure, Quaternary/drug effects , Protein Structure, Tertiary/drug effects , Temperature , Thermodynamics
18.
J Biol Chem ; 276(4): 2742-51, 2001 Jan 26.
Article in English | MEDLINE | ID: mdl-11056169

ABSTRACT

We have studied the biochemical features, the conformational preferences in solution, and the DNA binding properties of human p8 (hp8), a nucleoprotein whose expression is affected during acute pancreatitis. Biochemical studies show that hp8 has properties of the high mobility group proteins, HMG-I/Y. Structural studies have been carried out by using circular dichroism (near- and far-ultraviolet), Fourier transform infrared, and NMR spectroscopies. All the biophysical probes indicate that hp8 is monomeric (up to 1 mm concentration) and partially unfolded in solution. The protein seems to bind DNA weakly, as shown by electrophoretic gel shift studies. On the other hand, hp8 is a substrate for protein kinase A (PKA). The phosphorylated hp8 (PKAhp8) has a higher content of secondary structure than the nonphosphorylated protein, as concluded by Fourier transform infrared studies. PKAhp8 binds DNA strongly, as shown by the changes in circular dichroism spectra, and gel shift analysis. Thus, although there is not a high sequence homology with HMG-I/Y proteins, hp8 can be considered as a HMG-I/Y-like protein.


Subject(s)
DNA-Binding Proteins/metabolism , Growth Substances/metabolism , High Mobility Group Proteins , Neoplasm Proteins , Transcription Factors , Amino Acid Sequence , Basic Helix-Loop-Helix Transcription Factors , Circular Dichroism , Conserved Sequence , Cyclic AMP-Dependent Protein Kinases/metabolism , DNA-Binding Proteins/genetics , Growth Substances/genetics , HMGA1a Protein , Humans , Molecular Sequence Data , Nuclear Magnetic Resonance, Biomolecular , Phosphorylation , Protein Binding , Protein Structure, Secondary , Protein Structure, Tertiary , Sequence Homology, Amino Acid , Spectroscopy, Fourier Transform Infrared
20.
Med. intensiva ; 18(1): 11-15, 2001. ilus
Article in Spanish | BINACIS | ID: bin-8684

ABSTRACT

La sepsis grave mantiene una elevada mortalidad en los últimos 40 años. Múltiples intentos por moderar la respuesta inflamatoria sistémica, mediante la infusión de inmunoglobulinas (Ig), han demostrado resultados controvertidos. Los diferentes criterios de diagnóstico, la diferente población en estudio y reducido número de pacientes, dificultan obtener conclusiones definitivas. Objetivo: Evaluar si la infusión de Ig mejora la evolución de pacientes con sepsis abdominal. Lugar: 7 unidades de Cuidados Intensivos Polivalentes. Diseño: Estudio multicéntrico, prospectivo, randomizado, doble ciego. Intervenciones: Los pacientes fueron asignados aleatoriamente para recibir Ig (Grupo A: GA) o placebo (Grupo B: GB) conjuntamente con el tratamiento habitual de la sepsis abdominal. Resultado: Treinta y siete pacientes fueron incluidos en el estudio. 20 en el GA y 17 en el GB. El nivel medio de APACHE II, TISS, las disfunciones y fallas orgánicas no fueron diferentes entre los grupos. La mortalidad general fue del 16,7 por ciento, superior en el GB (26 por ciento) respecto del GA (5 por ciento) aunque tal diferencia no alcanzó significación estadística. Conclusiones: Nuestros hallazgos sugieren, que la infusión de Ig (Pentaglobin) en pacientes con sepsis abdominal con foco resuelto, reduce la mortalidad al alta de la UCI. Nuevos estudios clínicos son necesarios para confirmar los presentes hallazgos (AU)


Subject(s)
Humans , Male , Adult , Female , Middle Aged , Aged , Immunoglobulins, Intravenous/therapeutic use , Sepsis/therapy , Prospective Studies , Multicenter Studies as Topic , Treatment Outcome , Immunoglobulins/therapeutic use , Clinical Trials as Topic , Sepsis/etiology , Pelvic Inflammatory Disease/therapy , Peritonitis/therapy , Cholangitis/therapy , Double-Blind Method
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