RESUMEN

Abnormal lipid composition and metabolism of plasma lipoproteins play a crucial role in the pathogenesis of coronary heart disease (CHD). A (1)H NMR-based lipidomic approach was used to investigate the correlation of coronary artery stenosis with the atherogenic (non-HDL) and atheroprotective (HDL) lipid profiles in 99 patients with CHD of various stages of disease and compared with 60 patients with normal coronary arteries (NCA), all documented in coronary angiography. The pattern recognition models created from lipid profiles predicted the presence of CHD with a sensitivity of 87% and a specificity of 88% in the HDL model and with 90% and 89% in the non-HDL model, respectively. Patients with mild, moderate, and severe coronary artery stenosis were progressively differentiated from those with NCA in the non-HDL model with a statistically significant separation of severe stage from both mild and moderate. In the HDL model, the progressive differentiation of the disease stages was statistically significant only between patients with mild and severe coronary artery stenosis. The lipid constituents of lipoproteins that mainly characterized the initial stages and then the progression of the disease were the high levels of saturated fatty acids in lipids in both HDL and non-HDL particles, the low levels of HDL-phosphatidylcholine, HDL-sphingomyelin, and omega-3 fatty acids and linoleic acid in lipids in non-HDL particles. The conventional lipid marker, total cholesterol, found in low levels in HDL and in high levels in non-HDL, also contributed to the onset of the disease but with a much lower coefficient of significance. (1)H NMR-based lipidomic analysis of atherogenic and atheroprotective lipoproteins could contribute to the early evaluation of the onset of coronary artery disease and possibly to the establishment of an appropriate therapeutic option.

Asunto(s)

Enfermedad Coronaria/sangre , Metabolismo de los Lípidos , Lipoproteínas/sangre , Lipoproteínas/metabolismo , Espectroscopía de Protones por Resonancia Magnética/métodos , Anciano , Aterosclerosis , Enfermedad Coronaria/patología , Vasos Coronarios/metabolismo , Progresión de la Enfermedad , Femenino , Humanos , Lipoproteínas HDL/sangre , Lipoproteínas HDL/metabolismo , Masculino , Persona de Mediana Edad , Reproducibilidad de los Resultados , Índice de Severidad de la Enfermedad

RESUMEN

A (1)H NMR-based lipid profiling approach was used to investigate the prediction of coronary heart disease (CHD) and examine the confounding effect of factors such as gender, triglycerides, HDL-cholesterol and age levels on the prediction of disease. The HDL and non-HDL lipid profiles in 47 patients with triple vessel disease (TVD) and 41 patients with normal coronary arteries (NCA) both documented angiographically were generated. The presence of CHD was predicted with a sensitivity and specificity of 52% and 75% for HDL model and 78% and 80% for non-HDL, respectively. The lipid constituents of HDL lipoproteins which contributed to the separation between the two groups were the saturated fatty acids, cholesterol, total omega-3 fatty acids, degree of unsaturation, diallylic protons from polyunsaturated fatty acids, linoleic acid and, to a lesser extent, the number of fatty acids, triglycerides, unsaturated fatty acids and phosphatidylcholine. Respectively, for non-HDL, lipoproteins were the saturated fatty acids, number of fatty acids, cholesterol, unsaturated fatty acids and phosphatidylcholine. Gender, triglycerides, HDL-cholesterol and age influenced the lipid constituents of HDL and non-HDL lipoproteins that contributed to the separation between subgroups and confounded the predictive power of the models. NMR-based lipid profiling analysis could contribute to the identification of noninvasive markers for the presence and the development of the disease.

Asunto(s)

HDL-Colesterol/sangre , Enfermedad Coronaria/sangre , Lípidos/sangre , Resonancia Magnética Nuclear Biomolecular/métodos , Anciano , Femenino , Humanos , Masculino , Persona de Mediana Edad , Triglicéridos/sangre

RESUMEN

DsbD transmembrane protein dispatches electrons to periplasmic Trx/DsbE-like partners via specific interactions with its N-terminal domain, nDsbD. In the present study, PilB N-terminal domain (NterPilB) is shown to efficiently accept electrons coming from nDsbD from Neisseria meningitidis. Using an NMR-driven docking approach, we have modeled the structure of a mixed disulfide complex between NterPilB and nDsbD. We show the needed opening of nDsbD cap-loop whereas NterPilB FLHE loop does not seem essential in the formation and stabilization of the complex. Relaxation analysis performed on backbone amide groups highlights a kind of dynamics transfer from nDsbD cap-loop on NterPilB alpha1 helix, suggesting that a mobility contribution is required not only for the formation of the mixed disulfide complex, but also for its disruption. Taking into account previous X-ray data on covalent complexes involving nDsbD, a cartoon of interactions between Trx-like partners and nDsbD is proposed that illustrates the adaptability of nDsbD.

Asunto(s)

Proteínas Bacterianas/química , Proteínas Bacterianas/metabolismo , Disulfuros/química , Disulfuros/metabolismo , Neisseria meningitidis/metabolismo , Oxidorreductasas/química , Oxidorreductasas/metabolismo , Proteínas Bacterianas/aislamiento & purificación , Sitios de Unión , Disulfuros/aislamiento & purificación , Electrones , Cinética , Proteínas de la Membrana/química , Proteínas de la Membrana/metabolismo , Modelos Moleculares , Resonancia Magnética Nuclear Biomolecular , Oxidorreductasas/aislamiento & purificación , Periplasma/metabolismo , Unión Proteica , Estructura Secundaria de Proteína , Estructura Terciaria de Proteína

RESUMEN

The phosphopeptide P140 issued from the spliceosomal U1-70K snRNP protein is recognized by lupus CD4(+) T cells, transiently abolishes T cell reactivity to other spliceosomal peptides in P140-treated MRL/lpr mice, and ameliorates their clinical features. P140 modulates lupus patients' T cell response ex vivo and is currently included in phase IIb clinical trials. Its underlying mechanism of action remains elusive. Here we show that P140 peptide binds a unique cell-surface receptor, the constitutively-expressed chaperone HSC70 protein, known as a presenting-protein. P140 induces apoptosis of activated MRL/lpr CD4(+) T cells. In P140-treated mice, it increases peripheral blood lymphocyte apoptosis and decreases B cell, activated T cell, and CD4(-)CD8(-)B220(+) T cell counts via a specific mechanism strictly depending on gammadelta T cells. Expression of inflammation-linked genes is rapidly regulated in CD4(+) T cells. This work led us to identify a powerful pathway taken by a newly-designed therapeutic peptide to immunomodulate lupus autoimmunity.

Asunto(s)

Proteínas del Choque Térmico HSC70/metabolismo , Lupus Eritematoso Sistémico/inmunología , Fragmentos de Péptidos/farmacología , Linfocitos T Reguladores/inmunología , Animales , Apoptosis , Linfocitos B/metabolismo , Sitios de Unión , Regulación hacia Abajo , Técnica del Anticuerpo Fluorescente , Lupus Eritematoso Sistémico/metabolismo , Lupus Eritematoso Sistémico/terapia , Ratones , Ratones Endogámicos MRL lpr , Modelos Biológicos , Fragmentos de Péptidos/química , Fragmentos de Péptidos/metabolismo , Receptores de Antígenos de Linfocitos T gamma-delta/análisis , Ribonucleoproteína Nuclear Pequeña U1/inmunología , Ribonucleoproteína Nuclear Pequeña U1/metabolismo , Resonancia por Plasmón de Superficie , Linfocitos T/inmunología

RESUMEN

Human beta1,4-GalT (galactosyltransferase)7 is involved in the biosynthesis of the tetrasaccharide linker protein region (GlcAbeta1-->3Galbeta1-->3Galbeta1-->4Xylbeta1) (where GlcA is glucuronic acid and Xyl is xylose) of proteoglycans, by catalysing the transfer of Gal (galactose) from the uridine 5'-diphosphogalactose to a Xyl residue. This reaction is rate-limiting in glycosaminoglycan biosynthesis. In the present study, we established a large-scale production system of beta1,4-GalT7 fused with the maltose-binding protein to study substrate recognition. Calorimetric binding studies showed that the binding of the donor substrate UDP-Gal largely promoted binding of the acceptor substrate. To identify the structural basis governing substrate recognition, we used a fragment-based approach involving the artificial breakdown of the donor substrate into smaller fragments and characterization of their respective binding to the enzyme by isothermal titration calorimetry. The beta-phosphate, and to a lesser extent the alpha-phosphate, largely contributed to the binding energy. However, the uridine moiety was found to be essential for the optimal positioning of the donor substrate within the binding site. Unexpectedly, the contribution of the Gal moiety in substrate recognition was found to be negligible. Indeed, UDP-Gal, but also various UDP-sugars, could bind to beta1,4-GalT7. Surprisingly, in contrast with other GalTs, soluble beta1,4-GalT7 was able to transfer Glc (glucose), Xyl and, to a lesser extent GlcA and GlcNAc (N-acetyl glucosamine), to acceptor sugars, whereas UDP-Man (mannose) and UDP-GalNAc (N-acetyl galactosamine) were not substrates.

Asunto(s)

Galactosiltransferasas/metabolismo , Proteínas Portadoras/genética , Escherichia coli/enzimología , Galactosiltransferasas/antagonistas & inhibidores , Galactosiltransferasas/química , Galactosiltransferasas/aislamiento & purificación , Células HeLa , Humanos , Cinética , Proteínas de Unión a Maltosa , Resonancia Magnética Nuclear Biomolecular , Proteínas Recombinantes de Fusión/aislamiento & purificación , Especificidad por Sustrato , Termodinámica , Azúcares de Uridina Difosfato/metabolismo

RESUMEN

The DsbD protein is essential for electron transfer from the cytoplasm to the periplasm of Gram-negative bacteria. Its N-terminal domain dispatches electrons coming from cytoplasmic thioredoxin (Trx), via its central transmembrane and C-terminal domains, to its periplasmic partners: DsbC, DsbE/CcmG, and DsbG. Previous structural studies described the latter proteins as Trx-like folds possessing a characteristic C-X-X-C motif able to generate a disulfide bond upon oxidation. The Escherichia coli nDsbD displays an immunoglobulin-like fold in which two cysteine residues (Cys103 and Cys109) allow a disulfide bond exchange with its biological partners.We have determined the structure in solution and the backbone dynamics of the C103S mutant of the N-terminal domain of DsbD from Neisseria meningitidis. Our results highlight significant structural changes concerning the beta-sheets and the local topology of the active site compared with the oxidized form of the E. coli nDsbD. The structure reveals a "cap loop" covering the active site, similar to the oxidized E. coli nDsbD X-ray structure. However, regions featuring enhanced mobility were observed both near to and distant from the active site, revealing a capacity of structural adjustments in the active site and in putative interaction areas with nDsbD biological partners. Results are discussed in terms of functional consequences.

Asunto(s)

Cisteína/genética , Proteínas Mutantes/química , Mutación/genética , Neisseria meningitidis/enzimología , Oxidorreductasas/química , Oxidorreductasas/genética , Serina/genética , Dominio Catalítico , Espectroscopía de Resonancia Magnética , Modelos Moleculares , Proteínas Mutantes/genética , Estructura Terciaria de Proteína/genética , Soluciones

RESUMEN

The secreted form of the PilB protein was proposed to be involved in pathogen survival fighting against the defensive host's oxidative burst. PilB protein is composed of three domains. The central and the C-terminal domains display methionine sulfoxide reductase A and B activities, respectively. The N-terminal domain, which possesses a CXXC motif, was recently shown to regenerate in vitro the reduced forms of the methionine sulfoxide reductase domains of PilB from their oxidized forms, as does the thioredoxin 1 from E. coli, via a disulfide bond exchange. The thioredoxin-like N-terminal domain belongs to the cytochrome maturation protein structural family, but it possesses a unique additional segment (99)FLHE (102) localized in a loop. This segment covers one edge of the active site in the crystal structure of the reduced form of the N-terminal domain of PilB. We have determined the solution structure and the dynamics of the N-terminal domain from Neisseria meningitidis, in its reduced and oxidized forms. The FLHE loop adopts, in both redox states, a well-defined conformation. Subtle conformational and dynamic changes upon oxidation are highlighted around the active site, as well as in the FLHE loop. The functional consequences of the cytochrome maturation protein topology and those of the presence of FLHE loop are discussed in relation to the enzymatic properties of the N-terminal domain.

Asunto(s)

Proteínas Bacterianas/química , Proteínas Bacterianas/metabolismo , Neisseria meningitidis/metabolismo , Oxidorreductasas/química , Oxidorreductasas/metabolismo , Concentración de Iones de Hidrógeno , Espectroscopía de Resonancia Magnética , Modelos Moleculares , Oxidación-Reducción , Conformación Proteica , Estructura Terciaria de Proteína

RESUMEN

INTRODUCTION: Alterations in the lipid composition and overall structure of plasma lipoproteins have been correlated with pathological situations such as dyslipidaemia, coronary heart disease (CHD), hypertension, and renal disease. In the present study 1H NMR spectroscopy was used to analyse the lipid composition of HDL and nonHDL lipoproteins in patients with triple vessel CHD and in patients with normal coronary arteries. METHODS: Serum samples were collected from 50 patients with triple vessel CHD and 41 patients with normal coronary vessels, both documented angiographically. The classical risk factors for CHD were recorded and each patient's standard lipid profile was determined. HDL and nonHDL lipoprotein particles were separated by precipitation with Dextran Sulphate/MgCl2. HDL and nonHDL lipid fractions were extracted with chloroform:methanol (1:2, v/v). 1H NMR spectra were recorded on a Bruker DRX-600 spectrometer. RESULTS: In the HDL fraction of patients with triple vessel disease the percentage of triglycerides was significantly higher than in those with normal coronary arteries, whereas the percentages of cholesterol esters, phosphatidylcholine and sphingomyelin, as well as polyunsaturated fatty acids, such as linoleic, arachidonic, and eicosapentaenoic, were significantly lower. In the nonHDL fraction significantly higher levels of triglycerides and lower levels of polyunsaturated fatty acids were observed. CONCLUSIONS: Patients with established CHD show significant alterations in the composition of plasma lipoproteins compared to those with normal coronary arteries. Further study of plasma lipoprotein composition might be able to identify components as indexes for the existence of CHD.

Asunto(s)

Enfermedad Coronaria/sangre , Lipoproteínas/sangre , Anciano , Estudios de Casos y Controles , Colesterol/sangre , Enfermedad Coronaria/diagnóstico , Enfermedad Coronaria/etiología , Ácidos Grasos/sangre , Femenino , Humanos , Espectroscopía de Resonancia Magnética , Masculino , Persona de Mediana Edad , Reproducibilidad de los Resultados , Factores de Riesgo

RESUMEN

We report the nearly complete 1H, 13C, and 15N resonance assignments of the C103S mutant of the N-terminal domain of DsbD from Neisseria meningitides. Secondary structure determination using CSI method leads to the prediction of nine beta-sheet parts.

Asunto(s)

Antígenos Bacterianos/química , Proteínas Bacterianas/química , Espectroscopía de Resonancia Magnética/métodos , Neisseria meningitidis/metabolismo , Secuencia de Aminoácidos , Isótopos de Carbono/química , Datos de Secuencia Molecular , Peso Molecular , Mutagénesis Sitio-Dirigida , Neisseria meningitidis/genética , Isótopos de Nitrógeno/química , Protones

RESUMEN

We report the nearly complete 1H, 13C and 15N resonance assignments of the oxidized form (Cys(67)-Cys(70)) of the N-terminal domain of PilB from Neisseria meningitidis. Secondary structure determination using CSI method and TALOS leads mainly to the prediction of 7 alpha-helical and 5 beta-sheet parts.

Asunto(s)

Proteínas Bacterianas/química , Neisseria meningitidis/química , Oxidorreductasas/química , Proteínas Bacterianas/genética , Cistina/química , Estructura Molecular , Neisseria meningitidis/genética , Resonancia Magnética Nuclear Biomolecular , Oxidación-Reducción , Oxidorreductasas/genética , Estructura Secundaria de Proteína , Estructura Terciaria de Proteína , Proteínas Recombinantes/química , Proteínas Recombinantes/genética

RESUMEN

Aiming at contributing to the development of potential atheroprotective agents, we report on the concept and design of two peptide models, which mimic the amphipathic helices of apoA-I and incorporate Met into their sequences to validate its role as oxidant scavenger: Ac-ESK(Palm)KELSKSW(10)SEM(13)LKEK(Palm)SKS-NH(2) (model 1 [W(10), M(13)]) and Ac-ESK(Palm)KELSKSM(10)SEW(13)LKEK(Palm)SKS-NH(2) (model 2 [M(10), W(13)]). Hydrophobic residues of both models cover about the half of the surface, while the positively and negatively charged residues constitute two separate clusters on the hydrophilic face. Palmitoyl groups were introduced into the Lys-N(epsilon)H(2) groups at positions 3 and 17 to contribute to the amphipathic character of the peptides and stabilize the nonpolar face of the helix. Conformational study by the combined application of 2D-NMR and molecular dynamics simulations, CD, FTIR, and fluorescence spectroscopy revealed that model 1 adopts helical conformation and Met is well exposed to the microenvironment. Model 2 that derives from model 1 by exchanging W(10) (model 1) with M(10) and M(13) (model 1) with W(13) also displays helical characteristics, while Met is rather shielded. Oxidation experiments indicated that model 1 exhibits a 2-fold more potent antioxidant activity towards LDL oxidation, compared to model 2, confirming the role of Met, when is devoid of steric hindrances, as oxidant scavenger for the protection of LDL.

Asunto(s)

Apolipoproteína A-I/química , Aterosclerosis/prevención & control , Péptidos/química , Péptidos/farmacología , Secuencia de Aminoácidos , Antioxidantes/síntesis química , Antioxidantes/química , Antioxidantes/farmacología , Dicroismo Circular , Diseño de Fármacos , Humanos , Técnicas In Vitro , Modelos Moleculares , Datos de Secuencia Molecular , Resonancia Magnética Nuclear Biomolecular , Péptidos/síntesis química , Conformación Proteica , Estructura Secundaria de Proteína , Espectrometría de Fluorescencia , Espectroscopía Infrarroja por Transformada de Fourier , Termodinámica , Triptófano/química

RESUMEN

Methionine sulfoxide reductases (Msr) reduce methionine sulfoxide (MetSO)-containing proteins, back to methionine (Met). MsrAs are stereospecific for the S epimer whereas MsrBs reduce the R epimer of MetSO. Although structurally unrelated, the Msrs characterized so far display a similar catalytic mechanism with formation of a sulfenic intermediate on the catalytic cysteine and a concomitant release of Met, followed by formation of at least one intramolecular disulfide bond (between the catalytic and a recycling cysteine), which is then reduced by thioredoxin. In the case of the MsrA from Escherichia coli, two disulfide bonds are formed, i.e. first between the catalytic Cys51 and the recycling Cys198 and then between Cys198 and the second recycling Cys206. Three crystal structures including E. coli and Mycobacterium tuberculosis MsrAs, which, for the latter, possesses only the unique recycling Cys198, have been solved so far. In these structures, the distances between the cysteine residues involved in the catalytic mechanism are too large to allow formation of the intramolecular disulfide bonds. Here structural and dynamical NMR studies of the reduced wild-type and the oxidized (Cys51-Cys198) forms of C86S/C206S MsrA from E. coli have been carried out. The mapping of MetSO substrate-bound C51A MsrA has also been performed. The data support (1) a conformational switch occurring subsequently to sulfenic acid formation and/or Met release that would be a prerequisite to form the Cys51-Cys198 bond and, (2) a high mobility of the C-terminal part of the Cys51-Cys198 oxidized form that would favor formation of the second Cys198-Cys206 disulfide bond.

Asunto(s)

Escherichia coli/enzimología , Oxidorreductasas/química , Sitios de Unión , Catálisis , Cristalografía por Rayos X , Escherichia coli/química , Espectroscopía de Resonancia Magnética , Metionina Sulfóxido Reductasas , Modelos Biológicos , Isótopos de Nitrógeno/química , Oxidación-Reducción , Estructura Terciaria de Proteína , Soluciones , Relación Estructura-Actividad , Especificidad por Sustrato

Asunto(s)

Proteínas de Escherichia coli/química , Oxidorreductasas/química , Isótopos de Carbono , Cisteína , Hidrógeno , Metionina Sulfóxido Reductasas , Isótopos de Nitrógeno , Resonancia Magnética Nuclear Biomolecular , Oxidación-Reducción , Conformación Proteica

RESUMEN

Synthetic carriers play an important role in immunogen presentation, due to their ability of inducing improved and specific responses to conjugated epitopes. Their influence on the bioactive conformation of the epitope, though admittedly crucial for relevant in vitro and in vivo applications, is difficult to evaluate, given the usual lack of information on the complex conformational features determined by the nature of the carrier and the mode of ligation. Using the Herpes simplex virus glycoprotein D-1 epitope (Leu(9)-Lys-Nle-Ala-Asp-Pro-Asn-Arg-Phe-Arg-Gly-Lys-Asp-Leu(22)) as a model, we have performed a detailed conformational analysis on the free epitope peptide in solution and on three constructs in which the epitope was conjugated to sequential oligopeptide carriers {Ac-[Lys-Aib-Gly](4)-OH (SOC(4))} (through either a thioether or an amide bond; Ac: acetyl) and polytuftsin oligomers {H-[Thr-Lys-Pro-Lys-Gly](4)-NH(2) (T20)}, (through a thioether bond). The analysis of the epitope conformation in the parent protein, in carrier-conjugated and free form, suggests that the beta-turn structure of the -Asp(13)-Pro-Asn-Arg(16)- segment is highly conserved and independent of the epitope form. However, small conformational variations were observed at the C-terminal part of the epitope, depending on the nature of the carrier.

Asunto(s)

Epítopos/análisis , Glicoproteínas/química , Herpesvirus Humano 1/química , Oligopéptidos/síntesis química , Secuencia de Aminoácidos , Sitios de Unión , Dicroismo Circular , Cristalización , Cristalografía por Rayos X , Glicoproteínas/inmunología , Herpesvirus Humano 1/inmunología , Enlace de Hidrógeno , Modelos Moleculares , Datos de Secuencia Molecular , Resonancia Magnética Nuclear Biomolecular , Oligopéptidos/inmunología , Unión Proteica , Conformación Proteica , Estructura Secundaria de Proteína , Trifluoroetanol/química , Agua/química

RESUMEN

La/SSB phosphoprotein is the target antigen of autoantibodies in sera of patients with Sjögren's syndrome (SS) and systemic lupus erythematosus (SLE). Among other structural and function motifs, four phosphorylation sites are encompassed in the primary sequence of La/SSB. Two of them (Thr-362 and Ser-366) are located within GSGKGKVQFQGKKTKFASDD (346-368) and one (Thr-302) within VTWEVLEGEVEKEALKKI (301-318), which are main B-cell epitopes of La/SSB. With the aim to investigate how phosphorylation, one of the most common posttranslational protein modifications, affects the antigenic and conformational characteristics of the La/SSB epitopes, we synthesized and studied the phosphorylated epitopes La/SSB(346-368)-P, La/SSB(359-368)-P, and La/SSB(301-318)-P with respect to their nonphosphorylated counterparts. Anti-La/SSB positive sera from SS and SLE patients are better recognized by the phosphorylated epitopes compared to their nonphosphorylated counterparts. Conformational analysis by (1)H nuclear magnetic resonance spectroscopy and molecular dynamics showed that the phosphorylated epitopes adopt different structural characteristics from those of the corresponding nonphosphorylated epitopes. It is concluded that phosphorylation can create neoepitopes with altered functions, compared to the nonphosphorylated epitopes, which might be seen from the immune system as "foreign."

Asunto(s)

Autoantígenos/inmunología , Epítopos/química , Ribonucleoproteínas/inmunología , Secuencia de Aminoácidos , Autoantígenos/química , Autoantígenos/clasificación , Simulación por Computador , Ensayo de Inmunoadsorción Enzimática , Epítopos/genética , Epítopos/inmunología , Células HeLa , Humanos , Enlace de Hidrógeno , Lupus Eritematoso Sistémico/sangre , Lupus Eritematoso Sistémico/inmunología , Espectrometría de Masas , Microscopía Confocal , Modelos Moleculares , Conformación Molecular , Peso Molecular , Resonancia Magnética Nuclear Biomolecular , Péptidos/genética , Péptidos/inmunología , Fosforilación , Ribonucleoproteínas/química , Ribonucleoproteínas/clasificación , Síndrome de Sjögren/sangre , Síndrome de Sjögren/inmunología , Espectrometría de Masa por Ionización de Electrospray , Antígeno SS-B

Asunto(s)

Proteínas Bacterianas/química , Neisseria meningitidis/enzimología , Oxidorreductasas/química , Isótopos de Carbono , Hidrógeno , Isótopos de Nitrógeno , Resonancia Magnética Nuclear Biomolecular , Fragmentos de Péptidos/química

RESUMEN

Assignment of heteronuclear and homonuclear multidimensional NMR spectra permits determination of the first three-dimensional solution structure of a higher-plant thioredoxin h. The collection of 1906 distance restraints, 137 TALOS-derived dihedral restraints, and 66 hydrogen bonds was used in the restrained molecular dynamics protocol to calculate the structure of the reduced form of thioredoxin h1 from poplar with an atomic rmsd of 0.60 +/- 0.12 A. This enzyme exhibits an unusual active site with the sequence WCPPC and original properties in terms of stability and specificity. Compared to other known thioredoxin structures, thioredoxin h1 from poplar adopts the classical "Trx fold". Its atypical active site possesses a conformation similar to that of other common thioredoxins but appears to be more rigid. Moreover, the hydrogen bond network, stabilizing the in-core beta-sheet, is tighter than in Chlamydomonas reinhardtii, explaining the difference in thermostability.

Asunto(s)

Proteínas de Plantas/química , Proteínas de Plantas/metabolismo , Populus , Tiorredoxinas/química , Tiorredoxinas/metabolismo , Secuencia de Aminoácidos , Aminoácidos Aromáticos/química , Aminoácidos Aromáticos/metabolismo , Sitios de Unión , Isótopos de Carbono/metabolismo , Cristalografía por Rayos X , Enlace de Hidrógeno , Datos de Secuencia Molecular , Isótopos de Nitrógeno/metabolismo , Resonancia Magnética Nuclear Biomolecular/métodos , Oxidación-Reducción , Estructura Secundaria de Proteína , Protones , Soluciones , Termodinámica , Tiorredoxina h

Asunto(s)

Neisseria meningitidis/enzimología , Oxidorreductasas/química , Secuencia de Aminoácidos , Isótopos de Carbono , Metionina Sulfóxido Reductasas , Datos de Secuencia Molecular , Isótopos de Nitrógeno , Resonancia Magnética Nuclear Biomolecular , Estructura Secundaria de Proteína , Protones , Alineación de Secuencia

Asunto(s)

Populus/química , Tiorredoxinas/química , Tiorredoxinas/metabolismo , Secuencia de Aminoácidos , Sitios de Unión , Isótopos de Carbono , Datos de Secuencia Molecular , Isótopos de Nitrógeno , Resonancia Magnética Nuclear Biomolecular , Oxidación-Reducción , Protones , Tiorredoxina h

RESUMEN

Application of complementary B and T cell epitopes in inducing anti-idiotypic and anti-clonotypic antibodies capable of regulating or suppressing the autoimmune responses in experimental autoimmune myasthenia gravis (EAMG), allergic neuritis (EAN) and allergic encephalomyelitis (EAE) has been the stimulus of many research efforts. Studies on the idiotypic/anti-idiotypic network of anti-La/SSB positive sera from patients with Sjogren's Syndrome (SS) and Systemic Lupus Erythematosus (SLE) and on animals immunized with the complementary epitopes are presented.

Asunto(s)

Anticuerpos Antiidiotipos/inmunología , Enfermedades Autoinmunes/inmunología , Epítopos/inmunología , Péptidos/inmunología , Secuencia de Aminoácidos , Linfocitos B/inmunología , Encefalomielitis Autoinmune Experimental/inmunología , Epítopos/química , Lupus Eritematoso Sistémico/inmunología , Modelos Moleculares , Datos de Secuencia Molecular , Miastenia Gravis/inmunología , Neuritis Autoinmune Experimental/inmunología , Péptidos/química , Síndrome de Sjögren/inmunología , Espectrometría de Masa por Ionización de Electrospray , Resonancia por Plasmón de Superficie , Linfocitos T/inmunología