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1.
Brain Res ; 1459: 71-80, 2012 Jun 12.
Artículo en Inglés | MEDLINE | ID: mdl-22560502

RESUMEN

α-Synuclein is a neuronal protein implicated in the etiology of Parkinson's disease (PD) and dementia with Lewy bodies (DLB). Whilst increased α-synuclein expression due to gene duplication or triplication can cause familial PD, previous studies of α-synuclein levels in idiopathic disease have produced conflicting data. We quantified α-synuclein mRNA and soluble protein in five human post-mortem brain regions from four groups of individuals with PD, DLB, Alzheimer's disease (AD) and matched controls. α-Synuclein mRNA levels, measured using quantitative real-time PCR, did not differ significantly between groups in any brain regions examined. In contrast, levels of soluble α-synuclein protein, measured by ELISA, were significantly lower in 4 of the 5 regions for patients with DLB, and in 2 of the 5 regions for patients with PD, compared to controls. Soluble α-synuclein protein levels were not significantly different in the AD patients, compared to controls, in 4 of the 5 regions. This study indicates that although levels of soluble α-synuclein protein are lower in DLB and PD, there is no evidence for a corresponding decrease in α-synuclein mRNA levels. This might result from altered translation, or removal of α-synuclein protein from a soluble detectable state, either by turnover or conversion to an insoluble form.


Asunto(s)
Enfermedad de Alzheimer/patología , Encéfalo/metabolismo , Enfermedad por Cuerpos de Lewy/patología , Enfermedad de Parkinson/patología , ARN Mensajero/metabolismo , alfa-Sinucleína/genética , alfa-Sinucleína/metabolismo , Anciano , Anciano de 80 o más Años , Encéfalo/patología , Ensayo de Inmunoadsorción Enzimática , Femenino , Regulación de la Expresión Génica/fisiología , Humanos , Masculino , Cambios Post Mortem
2.
J Alzheimers Dis ; 29(4): 863-73, 2012.
Artículo en Inglés | MEDLINE | ID: mdl-22349685

RESUMEN

Zinc (Zn2+) is concentrated into pre-synaptic vesicles and co-released with neurotransmitter at some synapses. Zn2+ can accelerate assembly of the amyloid-ß peptides (Aß) and tau protein central to the neuropathological changes found in Alzheimer's disease (AD). Altered protein levels of the membrane Zn2+ transporters ZnT1, ZnT4, and ZnT6 have been reported in AD postmortem brain tissue. The present study analyzed mRNA levels of five established (LIV1, ZIP1, ZnT1, ZnT4, and ZnT6) and one potential (PRNP) Zn2+ transporter in human postmortem brain tissue from Braak-staged individuals with AD and controls using quantitative real-time PCR. Four cortical regions (middle temporal gyrus, superior occipital gyrus, superior parietal gyrus, and superior frontal gyrus) and cerebellum were examined. PRNP mRNA levels were decreased by ∼30% in all four cortical regions examined in AD patients, but unchanged in the cerebellum. In contrast, some increases in mRNA levels of the other more established Zn2+ transporters (LIV1, ZIP1, ZnT1, ZnT6) were found in AD cortex. The ratios of the mRNA levels of LIV1, ZIP1, ZnT1, ZnT4, and ZnT6/mRNA level of neuron specific enolase increased significantly as the disease progressed and Braak stage increased. Significant correlations were also identified between mRNA levels of several of the Zn2+ transporters investigated. These expression changes could either reflect or cause the altered cortical Zn2+ distribution in AD, potentially increasing the likelihood of interactions between Zn2+ and Aß or tau protein.


Asunto(s)
Enfermedad de Alzheimer/patología , Encéfalo/metabolismo , Proteínas Portadoras/genética , ARN Mensajero/metabolismo , Anciano , Anciano de 80 o más Años , Análisis de Varianza , Encéfalo/patología , Proteínas Portadoras/clasificación , Proteínas Portadoras/metabolismo , Estudios de Casos y Controles , Proteínas de Transporte de Catión/genética , Proteínas de Transporte de Catión/metabolismo , Femenino , Proteína Ácida Fibrilar de la Glía/genética , Proteína Ácida Fibrilar de la Glía/metabolismo , Humanos , Masculino , Proteínas de Neoplasias/genética , Proteínas de Neoplasias/metabolismo , Fosfopiruvato Hidratasa/genética , Fosfopiruvato Hidratasa/metabolismo , Cambios Post Mortem , Estadística como Asunto , Estadísticas no Paramétricas
3.
J Alzheimers Dis ; 22(4): 1111-22, 2010.
Artículo en Inglés | MEDLINE | ID: mdl-20930286

RESUMEN

ß-site AßPP cleaving enzyme 1 (BACE1) catalyses the rate-limiting step for production of amyloid-ß (Aß) peptides, involved in the pathological cascade underlying Alzheimer's disease (AD). Elevated BACE1 protein levels and activity have been reported in AD postmortem brains. Our study explored whether this was due to elevated BACE1 mRNA expression. RNA was prepared from five brain regions in three study groups: controls, individuals with AD, and another neurodegenerative disease group affected by either Parkinson's disease (PD) or dementia with Lewy bodies (DLB). BACE1 mRNA levels were measured using quantitative realtime PCR (qPCR) and analyzed by qbasePLUS using validated stably-expressed reference genes. Expression of glial and neuronal markers (glial fibrillary acidic protein (GFAP) and neuron-specific enolase (NSE), respectively) were also analyzed to quantify the changing activities of these cell populations in the tissue. BACE1 mRNA levels were significantly elevated in medial temporal and superior parietal gyri, compared to the PD/DLB and/or control groups. Superior frontal gryus BACE1 mRNA levels were significantly increased in the PD/DLB group, compared to AD and control groups. For the AD group, BACE1 mRNA changes were analyzed in the context of the reduced NSE mRNA, and strongly increased GFAP mRNA levels apparent as AD progressed (indicated by Braak stage). This analysis suggested that increased BACE1 mRNA expression in remaining neuronal cells may contribute to the increased BACE1 protein levels and activity found in brain regions affected by AD.


Asunto(s)
Enfermedad de Alzheimer/metabolismo , Secretasas de la Proteína Precursora del Amiloide/metabolismo , Ácido Aspártico Endopeptidasas/metabolismo , Encéfalo/metabolismo , Neuronas/metabolismo , Anciano , Anciano de 80 o más Años , Enfermedad de Alzheimer/genética , Secretasas de la Proteína Precursora del Amiloide/genética , Ácido Aspártico Endopeptidasas/genética , Femenino , Proteína Ácida Fibrilar de la Glía/genética , Proteína Ácida Fibrilar de la Glía/metabolismo , Humanos , Enfermedad por Cuerpos de Lewy/genética , Enfermedad por Cuerpos de Lewy/metabolismo , Masculino , Persona de Mediana Edad , Neuroglía/metabolismo , Enfermedad de Parkinson/genética , Enfermedad de Parkinson/metabolismo , Fosfopiruvato Hidratasa/genética , Fosfopiruvato Hidratasa/metabolismo , ARN Mensajero/genética , ARN Mensajero/metabolismo , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa
4.
Mol Neurodegener ; 4: 53, 2009 Dec 23.
Artículo en Inglés | MEDLINE | ID: mdl-20030848

RESUMEN

BACKGROUND: ZnT3 is a membrane Zn(2+ )transporter that is responsible for concentrating Zn(2+ )into neuronal presynaptic vesicles. Zn(2+ )homeostasis in the brain is relevant to Alzheimer's disease (AD) because Zn(2+ )released during neurotransmission may bind to Abeta peptides, accelerating the assembly of Abeta into oligomers which have been shown to impair synaptic function. RESULTS: We quantified ZnT3 mRNA levels in Braak-staged human post mortem (pm) brain tissue from medial temporal gyrus, superior occipital gyrus, superior parietal gyrus, superior frontal gyrus and cerebellum from individuals with AD (n = 28), and matched controls (n = 5) using quantitative real-time PCR. ZnT3 mRNA levels were significantly decreased in all four cortical regions examined in the AD patients, to 45-60% of control levels. This reduction was already apparent at Braak stage 4 in most cortical regions examined. Quantification of neuronal and glial-specific markers in the same samples (neuron-specific enolase, NSE; and glial fibrillary acidic protein, GFAP) indicated that loss of cortical ZnT3 expression was more pronounced, and occurred prior to, significant loss of NSE expression in the tissue. Significant increases in cortical GFAP expression were apparent as the disease progressed. No gene expression changes were observed in the cerebellum, which is relatively spared of AD neuropathology. CONCLUSIONS: This first study to quantify ZnT3 mRNA levels in human pm brain tissue from individuals with AD and controls has revealed a significant loss of ZnT3 expression in cortical regions, suggesting that neuronal cells in particular show reduced expression of ZnT3 mRNA in the disease. This suggests that altered neuronal Zn(2+ )handling may be an early event in AD pathogenesis.

5.
Neuromolecular Med ; 11(4): 337-44, 2009.
Artículo en Inglés | MEDLINE | ID: mdl-19669607

RESUMEN

Beta-site amyloid precursor protein cleaving enzyme (BACE1) is the rate-limiting enzyme for production of beta-amyloid peptides (Abeta), which are proposed to drive the pathological changes found in Alzheimer's disease (AD). Reticulon 3 (RTN3) is a negative modulator of BACE1 (beta-secretase) proteolytic activity, while peptidylprolyl isomerase (cyclophilin)-like 2 (PPIL2) positively regulates BACE1 expression. The present study investigated whether there was any association between genetic variation in RTN3 and PPIL2, and either risk for AD, or levels of platelet beta-secretase activity, in a large Northern Irish case-control sample. Four hundred and sixty-nine patients with a diagnosis of probable AD (NINCDS-ADRDA criteria) and 347 control individuals (MMSE > 28/30) were genotyped. SNPs in both genes were selected by downloading genotype data from the International HapMap Project (Phase II) and tags selected using multimarker approach in Haploview, where r (2) > 0.8 and LOD > 3.0. Non-synonymous SNPs of interest were also included. Genotyping was performed by Sequenom iPLEX and TaqMan technologies. Alleles, genotypes and multi-marker haplotypes were tested for association with AD, and platelet beta-secretase activities were measured for a subset of individuals (n = 231). Eight SNPs in RTN3 and 7 in PPIL2 were genotyped. We found no significant associations between allele, genotype or haplotype frequencies and risk of AD. Further, there was no effect of genotype on platelet membrane beta-secretase activity. We conclude that common or potentially functional genetic variation in these BACE1 interacting proteins does not affect platelet membrane beta-secretase activity or contribute to risk of AD in this population.


Asunto(s)
Enfermedad de Alzheimer/enzimología , Enfermedad de Alzheimer/genética , Secretasas de la Proteína Precursora del Amiloide/metabolismo , Ácido Aspártico Endopeptidasas/metabolismo , Proteínas Portadoras/genética , Ciclofilinas/genética , Predisposición Genética a la Enfermedad , Proteínas de la Membrana/genética , Proteínas del Tejido Nervioso/genética , Anciano , Anciano de 80 o más Años , Plaquetas/enzimología , Femenino , Humanos , Masculino , Irlanda del Norte , Población Blanca/genética
6.
J Neurochem ; 108(2): 341-9, 2009 Jan.
Artículo en Inglés | MEDLINE | ID: mdl-19094065

RESUMEN

Research into the cause of Alzheimer's disease (AD) has identified strong connections to cholesterol. Cholesterol and cholesterol esters can modulate amyloid precursor protein (APP) processing, thus altering production of the Abeta peptides that deposit in cortical amyloid plaques. Processing depends on the encounter between APP and cellular secretases, and is thus subject to the influence of cholesterol-dependent factors including protein trafficking, and distribution between membrane subdomains. We have directly investigated endogenous membrane beta-secretase activity in the presence of a range of membrane cholesterol levels in SH-SY5Y human neuroblastoma cells and human platelets. Membrane cholesterol significantly influenced membrane beta-secretase activity in a biphasic manner, with positive correlations at higher membrane cholesterol levels, and negative correlations at lower membrane cholesterol levels. Platelets from individuals with AD or mild cognitive impairment (n = 172) were significantly more likely to lie within the negative correlation zone than control platelets (n = 171). Pharmacological inhibition of SH-SY5Y beta-secretase activity resulted in increased membrane cholesterol levels. Our findings are consistent with the existence of a homeostatic feedback loop between membrane cholesterol level and membrane beta-secretase activity, and suggest that this regulatory mechanism is disrupted in platelets from individuals with cognitive impairment.


Asunto(s)
Secretasas de la Proteína Precursora del Amiloide/metabolismo , Plaquetas/ultraestructura , Membrana Celular/metabolismo , Colesterol/metabolismo , Anciano , Anciano de 80 o más Años , Enfermedad de Alzheimer/sangre , Secretasas de la Proteína Precursora del Amiloide/antagonistas & inhibidores , Ácido Aspártico Endopeptidasas/metabolismo , Plaquetas/citología , Estudios de Casos y Controles , Línea Celular Tumoral , Membrana Celular/efectos de los fármacos , Colesterol/farmacología , Trastornos del Conocimiento/sangre , Medio de Cultivo Libre de Suero/farmacología , Relación Dosis-Respuesta a Droga , Inhibidores Enzimáticos , Ensayo de Inmunoadsorción Enzimática/métodos , Femenino , Humanos , Inhibidores de Hidroximetilglutaril-CoA Reductasas/farmacología , Modelos Lineales , Masculino , Neuroblastoma/patología , Neuroblastoma/ultraestructura , Estadísticas no Paramétricas , Fracciones Subcelulares , Factores de Tiempo , beta-Ciclodextrinas/farmacología
7.
Mol Cell Biochem ; 318(1-2): 43-51, 2008 Nov.
Artículo en Inglés | MEDLINE | ID: mdl-18587628

RESUMEN

IQGAPs are cytoskeletal scaffolding proteins which link signalling pathways to the reorganisation of actin and microtubules. Human IQGAP1 has four IQ motifs each of which binds to calmodulin. The same region has been implicated in binding to two calmodulin-like proteins, the myosin essential light chain Mlc1sa and the calcium and zinc ion binding protein S100B. Using synthetic peptides corresponding to the four IQ motifs of human IQGAP1, we showed by native gel electrophoresis that only the first IQ motif interacts with Mlc1sa. This IQ motif, and also the fourth, interacts with the budding yeast myosin essential light chain Mlc1p. The first and second IQ motifs interact with S100B in the presence of calcium ions. This clearly establishes that S100B can interact with its targets through IQ motifs in addition to interacting via previously reported sequences. These results are discussed in terms of the function of IQGAP1 and IQ motif recognition.


Asunto(s)
Cadenas Ligeras de Miosina/metabolismo , Factores de Crecimiento Nervioso/metabolismo , Proteínas S100/metabolismo , Proteínas Activadoras de ras GTPasa/química , Proteínas Activadoras de ras GTPasa/metabolismo , Secuencias de Aminoácidos , Secuencia de Aminoácidos , Humanos , Datos de Secuencia Molecular , Cadenas Ligeras de Miosina/aislamiento & purificación , Factores de Crecimiento Nervioso/aislamiento & purificación , Péptidos/metabolismo , Unión Proteica , Subunidad beta de la Proteína de Unión al Calcio S100 , Proteínas S100/aislamiento & purificación , Saccharomyces cerevisiae/metabolismo , Proteínas de Saccharomyces cerevisiae/metabolismo , Alineación de Secuencia , Relación Estructura-Actividad
8.
BMC Mol Biol ; 9: 46, 2008 May 06.
Artículo en Inglés | MEDLINE | ID: mdl-18460208

RESUMEN

BACKGROUND: Studies of gene expression in post mortem human brain can contribute to understanding of the pathophysiology of neurodegenerative diseases, including Alzheimer's disease (AD), Parkinson's disease (PD) and dementia with Lewy bodies (DLB). Quantitative real-time PCR (RT qPCR) is often used to analyse gene expression. The validity of results obtained using RT qPCR is reliant on accurate data normalization. Reference genes are generally used to normalize RT qPCR data. Given that expression of some commonly used reference genes is altered in certain conditions, this study aimed to establish which reference genes were stably expressed in post mortem brain tissue from individuals with AD, PD or DLB. RESULTS: The present study investigated the expression stability of 8 candidate reference genes, (ubiquitin C [UBC], tyrosine-3-monooxygenase [YWHAZ], RNA polymerase II polypeptide [RP II], hydroxymethylbilane synthase [HMBS], TATA box binding protein [TBP], beta-2-microglobulin [B2M], glyceraldehyde-3-phosphate dehydrogenase [GAPDH], and succinate dehydrogenase complex-subunit A, [SDHA]) in cerebellum and medial temporal gyrus of 6 AD, 6 PD, 6 DLB subjects, along with 5 matched controls using RT qPCR (TaqMan(R) Gene Expression Assays). Gene expression stability was analysed using geNorm to rank the candidate genes in order of decreasing stability in each disease group. The optimal number of genes recommended for accurate data normalization in each disease state was determined by pairwise variation analysis. CONCLUSION: This study identified validated sets of mRNAs which would be appropriate for the normalization of RT qPCR data when studying gene expression in brain tissue of AD, PD, DLB and control subjects.


Asunto(s)
Encéfalo/metabolismo , Perfilación de la Expresión Génica , Regulación de la Expresión Génica , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa/métodos , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa/normas , Humanos , ARN/metabolismo , Estándares de Referencia , Reproducibilidad de los Resultados , Programas Informáticos
9.
Mol Med ; 14(7-8): 451-64, 2008.
Artículo en Inglés | MEDLINE | ID: mdl-18368143

RESUMEN

Developing effective treatments for neurodegenerative diseases is one of the greatest medical challenges of the 21st century. Although many of these clinical entities have been recognized for more than a hundred years, it is only during the past twenty years that the molecular events that precipitate disease have begun to be understood. Protein aggregation is a common feature of many neurodegenerative diseases, and it is assumed that the aggregation process plays a central role in pathogenesis. In this process, one molecule (monomer) of a soluble protein interacts with other monomers of the same protein to form dimers, oligomers, and polymers. Conformation changes in three-dimensional structure of the protein, especially the formation of beta-strands, often accompany the process. Eventually, as the size of the aggregates increases, they may precipitate as insoluble amyloid fibrils, in which the structure is stabilized by the beta-strands interacting within a beta-sheet. In this review, we discuss this theme as it relates to the two most common neurodegenerative conditions-Alzheimer's and Parkinson's diseases.


Asunto(s)
Enfermedad de Alzheimer/etiología , Péptidos beta-Amiloides/metabolismo , Encéfalo/metabolismo , Enfermedad de Parkinson/etiología , Placa Amiloide/metabolismo , Enfermedad de Alzheimer/diagnóstico , Enfermedad de Alzheimer/epidemiología , Enfermedad de Alzheimer/metabolismo , Péptidos beta-Amiloides/fisiología , Péptidos beta-Amiloides/toxicidad , Encéfalo/patología , Humanos , Incidencia , Modelos Biológicos , Enfermedad de Parkinson/diagnóstico , Enfermedad de Parkinson/epidemiología , Enfermedad de Parkinson/metabolismo , alfa-Sinucleína/metabolismo , alfa-Sinucleína/fisiología
10.
Mech Ageing Dev ; 128(5-6): 378-82, 2007.
Artículo en Inglés | MEDLINE | ID: mdl-17531291

RESUMEN

Genetic variation of the alpha-synuclein gene (SNCA) is known to cause familial parkinsonism, however the role of SNCA variants in sporadic Parkinson's disease (PD) remains elusive. The present study identifies an association of common SNCA polymorphisms with disease susceptibility in a series of Irish PD patients. There is evidence for association with alternate regions, of protection and risk which may act independently/synergistically, within the promoter region (Rep1; OR: 0.59, 95% CI: 0.37-0.84) and the 3'UTR of the gene (rs356165; OR: 1.67, 95% CI: 1.08-2.58). Given previous reports of association a collaborative effort is required which may exploit global linkage disequilibrium patterns for SNCA and standardise polymorphic markers used in each population. It is now crucial to identify the susceptibility allele and elucidate its functionality which may generate a therapeutic target for PD.


Asunto(s)
Predisposición Genética a la Enfermedad , Enfermedad de Parkinson/genética , alfa-Sinucleína/genética , Anciano , Alelos , Femenino , Humanos , Masculino , Persona de Mediana Edad , Polimorfismo Genético
11.
Exp Brain Res ; 173(2): 223-33, 2006 Aug.
Artículo en Inglés | MEDLINE | ID: mdl-16733698

RESUMEN

An abundance of genetic, histopathological, and biochemical evidence has implicated the neuronal protein, alpha-synuclein (alpha-syn) as a key player in the development of several neurodegenerative diseases, the so-called synucleinopathies, of which Parkinson's disease (PD) is the most prevalent. Development of disease appears to be linked to events that increase the intracellular concentration of alpha-syn or cause its chemical modification, either of which can accelerate the rate at which it forms aggregates. Examples of such events include increased copy number of genes, decreased rate of degradation via the proteasome or other proteases, or altered forms of alpha-syn, such as truncations, missense mutations, or chemical modifications by oxidative reactions. Aggregated forms of the protein, especially newly formed soluble aggregates, are toxic to cells, so that one therapeutic strategy would be to reduce the rate at which such oligomerization occurs. We have therefore designed several peptides and also identified small molecules that can inhibit alpha-syn oligomerization and toxicity in vitro. These compounds could serve as lead compounds for the design of new drugs for the treatment of PD and related disorders in the future.


Asunto(s)
Antiparkinsonianos/farmacología , Antiparkinsonianos/uso terapéutico , Enfermedad de Parkinson/tratamiento farmacológico , alfa-Sinucleína/antagonistas & inhibidores , alfa-Sinucleína/toxicidad , Animales , Biomarcadores , Humanos , Cuerpos de Lewy/patología , Enfermedad de Parkinson/patología , alfa-Sinucleína/biosíntesis , Sinucleína beta/fisiología , gamma-Sinucleína/fisiología
12.
Parkinsonism Relat Disord ; 11(6): 349-52, 2005 Sep.
Artículo en Inglés | MEDLINE | ID: mdl-16102999

RESUMEN

The role of genetics in parkinsonism has been confirmed over the last decade with the identification of genetic variation in seven genes, which are causative in familial forms of the disorder. A number of pathogenic mutations have been identified in the latest gene LRRK2, with a Gly2019Ser amino acid substitution identified in two siblings and one patient with idiopathic Parkinson's disease from Ireland. The clinical features resemble the idiopathic variant with a tremor predominant clinical picture shared by the siblings, slow progression of symptoms, and no observation of cognitive disturbance in all. The family and the sporadic individual were apparently not related and originated from different regions of Ireland, although haplotype analysis does suggest they share a common founder. The influence of the G2019S substitution on protein function and disease phenotype has yet to be fully resolved, but its elucidation will undoubtedly further our understanding of the mechanisms underlying Parkinson's disease.


Asunto(s)
Enfermedad de Parkinson/genética , Enfermedad de Parkinson/fisiopatología , Proteínas Serina-Treonina Quinasas/genética , Adulto , Sustitución de Aminoácidos , Antiparkinsonianos/uso terapéutico , Cognición/fisiología , Progresión de la Enfermedad , Lateralidad Funcional/fisiología , Haplotipos , Humanos , Irlanda , Proteína 2 Quinasa Serina-Treonina Rica en Repeticiones de Leucina , Levodopa/uso terapéutico , Masculino , Persona de Mediana Edad , Mutación/fisiología , Pruebas Neuropsicológicas , Enfermedad de Parkinson/psicología
13.
Protein Pept Lett ; 11(3): 271-9, 2004 Jun.
Artículo en Inglés | MEDLINE | ID: mdl-15182228

RESUMEN

Alpha-synuclein is a major component of Lewy bodies in Parkinson's disease and is found associated with several other forms of dementia. As with other neurodegenerative diseases, the ability of alpha-synuclein to aggregate and form fibrillar deposits seems central to its pathology. We have defined a sequence within the NAC region of alpha-synuclein that is necessary for aggregation. Exploitation of chemically modified analogues of this peptide may produce inhibitors of aggregation.


Asunto(s)
Proteínas del Tejido Nervioso/química , Proteínas del Tejido Nervioso/metabolismo , Secuencia de Aminoácidos , Amiloidosis/tratamiento farmacológico , Amiloidosis/metabolismo , Animales , Humanos , Datos de Secuencia Molecular , Proteínas del Tejido Nervioso/clasificación , Enfermedad de Parkinson/metabolismo , Estructura Cuaternaria de Proteína , Sinucleínas , alfa-Sinucleína
14.
Neurosci Lett ; 359(1-2): 89-93, 2004 Apr 08.
Artículo en Inglés | MEDLINE | ID: mdl-15050719

RESUMEN

Alpha-synuclein has been linked to amyloidogenesis in Parkinson's disease and other neurodegenerative disorders. We have previously shown that a peptide comprising residues 68-78 of alpha-synuclein is the minimum fragment that, like alpha-synuclein itself, forms amyloid fibrils and exhibits toxicity towards cells in culture. Hughes et al. [J. Biol. Chem. 275 (2000) 25109] showed that an N-methylated derivative of Abeta(25-35) inhibited the formation of fibrils by Abeta(25-35) and reduced its toxicity. We have now extended this concept to an amyloidogenic alpha-synuclein-based peptide. Alpha-synuclein(68-78), N-methylated at G1y73, was compared to non-methylated peptide. Whereas alpha-synuclein(68-78) formed fibrils and was toxic to cells, the N-methylated analogue had neither of these properties. Moreover, an equimolar mixture of the non-methylated and methylated peptides formed very few fibrils and toxicity was markedly reduced.


Asunto(s)
Péptidos beta-Amiloides/toxicidad , Proteínas del Tejido Nervioso/toxicidad , Fragmentos de Péptidos/toxicidad , Placa Amiloide/efectos de los fármacos , Animales , Relación Dosis-Respuesta a Droga , Células PC12 , Placa Amiloide/patología , Ratas , Sinucleínas , alfa-Sinucleína
15.
J Biochem Biophys Methods ; 56(1-3): 233-42, 2003 Jun 30.
Artículo en Inglés | MEDLINE | ID: mdl-12834979

RESUMEN

Gel filtration on soft gels has been employed for over 40 years for the separation, desalting and molecular weight estimation of peptides and proteins. Technical improvements have given rise to high-performance size-exclusion chromatography (HPSEC) on rigid supports, giving more rapid run times and increased resolution. Initially, these packings were more suitable for the separation of proteins than of peptides, but supports that operate in the fractionation range <10,000 Daltons (Da) are now available. In this report, HPSEC is described in relation to its application to peptides, especially regarding purification, estimation of molecular weight and study of molecular associations.


Asunto(s)
Cromatografía en Gel/métodos , Geles/química , Péptidos/análisis , Péptidos/química , Dimerización , Sustancias Macromoleculares , Peso Molecular , Péptidos/aislamiento & purificación , Fotometría/métodos , Espectrometría de Masa por Ionización de Electrospray/métodos , Espectrofotometría/métodos
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