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1.
J Phys Chem B ; 123(31): 6694-6704, 2019 08 08.
Artigo em Inglês | MEDLINE | ID: mdl-31290327

RESUMO

Although successful in the structural determination of ordered biomolecules, the spectroscopic investigation of oligopeptides in solution is hindered by their complex and rapidly changing conformational ensemble. The measured circular dichroism (CD) spectrum of an oligopeptide is an ensemble average over all microstates, severely limiting its interpretation, in contrast to ordered biomolecules. Spectral deconvolution methods to estimate the secondary structure contributions in the ensemble are still mostly based on databases of larger ordered proteins. Here, we establish how the interpretation of CD spectra of oligopeptides can be enhanced by the ability to compute the same observable from a set of atomic coordinates. Focusing on two representative oligopeptides featuring a known propensity toward an α-helical and ß-hairpin motif, respectively, we compare and cross-validate the structural information coming from deconvolution of the experimental CD spectra, sequence-based de novo structure prediction, and molecular dynamics simulations based on enhanced sampling methods. We find that small conformational variations can give rise to significant changes in the CD signals. While for the simpler conformational landscape of the α-helical peptide de novo structure prediction can already give reasonable agreement with the experiment, an extended ensemble of conformers needs to be considered for the ß-hairpin sequence.


Assuntos
Oligopeptídeos/química , Sequência de Aminoácidos , Dicroísmo Circular , Análise por Conglomerados , Simulação de Dinâmica Molecular , Conformação Proteica em alfa-Hélice , Conformação Proteica em Folha beta
2.
Mutat Res Genet Toxicol Environ Mutagen ; 836(Pt A): 117-123, 2018 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-30389154

RESUMO

The ataxia telangiectasia mutated (ATM) protein is a pivotal multifunctional protein kinase predominantly involved in DNA damage response, as well as in maintaining overall functional integrity of the cells. Apart from playing its major role in regulating the cellular response to DNA damage, ATM, when mutated, can additionally determine oxidative stress, metabolic syndrome, mitochondrial dysfunction and neurodegeneration. In the present paper we aim to investigate the levels of oxidative stress potentially induced by the oxidizing rodent renal carcinogen KBrO3 in ATM-defective lymphoblastoid cell lines (LCLs) established from four classical AT patients (with different ATM mutations), one AT variant with reduced hypersensitivity to X rays, obligate AT heterozygotes and wild type intrafamilial control. A possible modulatory involvement of PARP in potentially induced oxidative stress is also evaluated following its inhibition with 3-aminobenzamide (3-AB). Treatments with KBrO3 clearly showed a marked hypersensitivity of the ATM-defective LCLs, including the AT variant. A marked and statistically significant reduction of KBrO3-induced chromosomal damage following inhibition of PARP by 3-AB, was observed in all AT LCLs, but not in those from the AT variant, AT heterozygotes and wild type intrafamilial control. This result is suggestive of a modulatory involvement of PARP in the hypersensitivity of ATM-defective cells to DNA oxidative damage.


Assuntos
Proteínas Mutadas de Ataxia Telangiectasia/deficiência , Bromatos/farmacologia , Dano ao DNA , Hipersensibilidade/tratamento farmacológico , Linfócitos/patologia , Poli(ADP-Ribose) Polimerase-1/antagonistas & inibidores , Inibidores de Poli(ADP-Ribose) Polimerases/farmacologia , Ataxia Telangiectasia/tratamento farmacológico , Ataxia Telangiectasia/enzimologia , Ataxia Telangiectasia/genética , Ataxia Telangiectasia/patologia , Células Cultivadas , Reparo do DNA , Humanos , Hipersensibilidade/genética , Hipersensibilidade/metabolismo , Hipersensibilidade/patologia , Linfócitos/efeitos dos fármacos , Linfócitos/efeitos da radiação , Estresse Oxidativo , Fosforilação
3.
Neuropediatrics ; 39(1): 43-5, 2008 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-18504682
4.
Neurology ; 66(8): 1207-10, 2006 Apr 25.
Artigo em Inglês | MEDLINE | ID: mdl-16636238

RESUMO

BACKGROUND: Ataxia with oculomotor apraxia type 2 (AOA2) is characterized by onset between age 10 and 22 years, cerebellar atrophy, peripheral neuropathy, oculomotor apraxia (OMA), and elevated serum alpha-fetoprotein (AFP) levels. Recessive mutations in SETX have been described in AOA2 patients. OBJECTIVE: To describe the clinical features of AOA2 and to identify the SETX mutations in 10 patients from four Italian families. METHODS: The patients underwent clinical examination, routine laboratory tests, nerve conduction studies, sural nerve biopsy, and brain MRI. All were screened for SETX mutations. RESULTS: All the patients had cerebellar features, including limb and truncal ataxia, and slurred speech. OMA was observed in two patients, extrapyramidal symptoms in two, and mental impairment in three. High serum AFP levels, motor and sensory axonal neuropathy, and marked cerebellar atrophy on MRI were detected in all the patients who underwent these examinations. Sural nerve biopsy revealed a severe depletion of large myelinated fibers in one patient, and both large and small myelinated fibers in another. Postmortem findings are also reported in one of the patients. Four different homozygous SETX mutations were found (a large-scale deletion, a missense change, a single-base deletion, and a splice-site mutation). CONCLUSIONS: The clinical phenotype of oculomotor apraxia type 2 is fairly homogeneous, showing only subtle intrafamilial variability. OMA is an inconstant finding. The identification of new mutations expands the array of SETX variants, and the finding of a missense change outside the helicase domain suggests the existence of at least one more functional region in the N-terminus of senataxin.


Assuntos
Apraxias/genética , Apraxias/patologia , Ataxia/genética , Ataxia/patologia , Doenças do Nervo Oculomotor/genética , Doenças do Nervo Oculomotor/patologia , Adolescente , Adulto , Idade de Início , Idoso , Apraxias/classificação , Apraxias/complicações , Ataxia/complicações , Atrofia , Cerebelo/patologia , Pré-Escolar , DNA Helicases , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Enzimas Multifuncionais , Mutação , Doenças do Nervo Oculomotor/complicações , Linhagem , Doenças do Sistema Nervoso Periférico/genética , Doenças do Sistema Nervoso Periférico/patologia , RNA Helicases/genética
5.
Anticancer Res ; 25(6C): 4451-4, 2005.
Artigo em Inglês | MEDLINE | ID: mdl-16334125

RESUMO

BACKGROUND: For many years surgery was the cornerstone of treatment for head and neck cancers and radiotherapy was the treatment of choice in adjuvant and advanced inoperable settings. Recently, induction sequential chemotherapy followed by radiotherapy has shown good tolerability and has prolonged the median overall survival. This phase II trial explored the feasibility of the concurrent association with radiotherapy of a full-dose chemotherapy based on an original schedule of docetaxel and cisplatin. PATIENTS AND METHODS: Twenty-four patients with head and neck squamous cell carcinoma (HNSCC) were enrolled. Taxotere (docetaxel) was administered on day 1, weekly for 6 weeks. The dose was 33 mg/m2 /w. Cisplatin was administered on day 2 at the dose of 70 mg/m2. Radiotherapy delivered was 60 Gy divided in 30 administrations over 6 weeks. RESULTS AND CONCLUSION: This schedule of treatment for HNSCC proved feasible. Appropriate support treatment, however, appears to be necessary for the feasibility of this concurrent chemo-radiotherapy.


Assuntos
Protocolos de Quimioterapia Combinada Antineoplásica/uso terapêutico , Carcinoma de Células Escamosas/tratamento farmacológico , Carcinoma de Células Escamosas/radioterapia , Neoplasias de Cabeça e Pescoço/tratamento farmacológico , Neoplasias de Cabeça e Pescoço/radioterapia , Idoso , Anemia/induzido quimicamente , Protocolos de Quimioterapia Combinada Antineoplásica/efeitos adversos , Cisplatino/administração & dosagem , Cisplatino/efeitos adversos , Terapia Combinada , Docetaxel , Estudos de Viabilidade , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Neutropenia/induzido quimicamente , Taxoides/administração & dosagem , Taxoides/efeitos adversos
6.
Cell Mol Life Sci ; 62(4): 485-91, 2005 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-15719174

RESUMO

The cells of an ataxia-oculomotor apraxia type 1 (AOA1) patient, homozygous for a new aprataxin mutation (T739C), were treated with camptothecin, an inhibitor of DNA topoisomerase I which induces DNA single-strand breaks. DNA damage was evaluated by cytogenetic analysis of chromosomal aberrations. The results obtained showed marked and dose-related increases in induced chromosomal aberrations in the patient and her heterozygous mother compared to the intrafamilial wild-type control. The alkaline comet assay confirmed this pattern. Moreover, the AOA1 cells did not show hypersensitivity to ionizing radiation, i.e. X-rays. These findings clearly indicate the direct involvement of aprataxin in the DNA single-strand-break repair machinery.


Assuntos
Apraxia Ideomotora/genética , Dano ao DNA , Reparo do DNA , Proteínas de Ligação a DNA/fisiologia , Proteínas Nucleares/fisiologia , Apraxia Ideomotora/diagnóstico , Camptotecina/farmacologia , Ataxia Cerebelar/diagnóstico , Ataxia Cerebelar/genética , Aberrações Cromossômicas , Ensaio Cometa , Dano ao DNA/genética , Reparo do DNA/genética , DNA Topoisomerases Tipo I/efeitos dos fármacos , DNA Topoisomerases Tipo I/fisiologia , DNA de Cadeia Simples/metabolismo , Proteínas de Ligação a DNA/análise , Proteínas de Ligação a DNA/genética , Diagnóstico Diferencial , Humanos , Masculino , Proteínas Nucleares/análise , Proteínas Nucleares/genética , Linhagem , Mutação Puntual/genética , Tolerância a Radiação/genética , Raios X
7.
Suppl Tumori ; 4(3): S215, 2005.
Artigo em Inglês | MEDLINE | ID: mdl-16437998

RESUMO

Usually head and neck cancer is treated with combined therapy, applying surgery, if possible, and then radiotherapy and chemotherapy in a sequential or concomitant way. Sequential approach seems to be preferred, because of the high toxicity rate of concomitant therapy. Platinum compounds and 5-fluorouracil are the standard drugs, but new drugs are entering therapeutic arena: gemcitabine and taxanes are the most promising ones. The efficacy of these drugs, especially in association with radiotherapy, must be assessed; moreover it is essential to ascertain how to associate these drugs to radiotherapy and to evaluate drug toxicity when combined with the latter. End point of the study here presented is a preliminar assessment of toxicity and feasibility of concurrent radio-chemoterapy with docetaxel and cisplatinum in patients with head and neck cancer. The number of enrolled patients and the relatively short time of follow up do not allow to evaluate treatment efficacy.


Assuntos
Protocolos de Quimioterapia Combinada Antineoplásica/uso terapêutico , Neoplasias de Cabeça e Pescoço/tratamento farmacológico , Neoplasias de Cabeça e Pescoço/radioterapia , Recidiva Local de Neoplasia/tratamento farmacológico , Recidiva Local de Neoplasia/radioterapia , Idoso , Antineoplásicos Fitogênicos/administração & dosagem , Cisplatino/administração & dosagem , Terapia Combinada , Docetaxel , Humanos , Pessoa de Meia-Idade , Estudos Prospectivos , Taxoides/administração & dosagem
8.
Neuropediatrics ; 32(5): 250-5, 2001 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-11748496

RESUMO

Progressive multifocal leukoencephalopathy (PML) is a fatal demyelinating disease due to infection with polyomavirus JC (JCV). PML occurs almost exclusively in immunocompromised patients, and although it has increased markedly in relation to AIDS, remains exceptional in children. We present the case of an immunocompromised child with hyperimmunoglobulin E recurrent infection syndrome (HIES) and pathologically-proven PML. HIES is a rare congenital immunodeficiency that to our knowledge has never before been reported in association with neurological complications. Following a recurrence of bronchopneumonia, the child's motor and cognitive functions deteriorated progressively in parallel with alterations on cerebral MRI. The neurological onset coincided with lymphocyte subset changes. PCR for JCV DNA did not detect the virus in CSF, and brain biopsy was required to secure the diagnosis. Antiviral treatment with cidofovir produced no benefit. Autopsy revealed the typical neuropathological findings of PML which were associated with inflammatory eosinophilic infiltrate (a marker of HIES). In accordance with the few pediatric PML cases reported and here reviewed, the child died five months after neurological onset.


Assuntos
Hipergamaglobulinemia/diagnóstico , Imunoglobulina E/sangue , Leucoencefalopatia Multifocal Progressiva/diagnóstico , Infecções Oportunistas/diagnóstico , Infecções Estafilocócicas/diagnóstico , Encéfalo/patologia , Criança , Encefalite/diagnóstico , Encefalite/patologia , Evolução Fatal , Humanos , Hipergamaglobulinemia/patologia , Leucoencefalopatia Multifocal Progressiva/patologia , Imageamento por Ressonância Magnética , Masculino , Oligodendroglia/patologia , Infecções Oportunistas/patologia , Recidiva , Infecções Estafilocócicas/patologia
9.
Hum Mol Genet ; 10(11): 1155-62, 2001 May 15.
Artigo em Inglês | MEDLINE | ID: mdl-11371508

RESUMO

We showed recently that mutation of the hMRE11 gene identified a new ataxia telangiectasia-like disorder (ATLD). In this report we describe the genomic organization of the hMRE11 gene and the analysis of a promoter region that appears to direct the divergent transcription of hMRE11 and the adjacent gene. The characterization of the genomic organization of the hMRE11 gene allowed us to determine the basis of an apparent null hMRE11 allele present in the mother and two patients in one of our two ATLD families. Polymorphic markers in the hMRE11 gene, including the promoter region, provided evidence that the mutated maternal allele was not deleted. An exon by exon search revealed the presence of a missense mutation in exon 15, the effect of which was to create a premature termination codon. Transcripts derived from the mutant allele were found to be subject to nonsense-mediated mRNA decay (NMD). Therefore, this allele was effectively null, because little if any mRNA from it was available for translation. The ATLD patients carrying this protein-truncating hMRE11 mutation have survived because the null allele they inherited from their mother is present with a missense mutation inherited from their father, which is expressed as normal levels of partially functional MRE11 protein. The mutation in the maternal hMRE11 allele of family 2 was also identified in a further unrelated Italian family with ATLD and also found to be subject to NMD.


Assuntos
Ataxia Telangiectasia/genética , Proteínas de Ligação a DNA/genética , Genoma , Mutação , Regiões Promotoras Genéticas , RNA Mensageiro/genética , Alelos , Ataxia Telangiectasia/metabolismo , Sequência de Bases , Linhagem Celular , Análise Mutacional de DNA , Proteínas de Ligação a DNA/metabolismo , Éxons , Vetores Genéticos , Humanos , Proteína Homóloga a MRE11 , Dados de Sequência Molecular , Biossíntese de Proteínas , Pseudogenes
10.
Prenat Diagn ; 19(6): 542-5, 1999 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-10416970

RESUMO

Ataxia telangiectasia (AT) is a severe autosomal recessive disease, rare but not infrequent in Italy. Owing to the seriousness of the disease, prenatal diagnosis has been attempted in the past by means of cytogenetic, biochemical, radio-biological and indirect molecular analyses. We performed the first direct molecular prenatal diagnosis of AT on a chorionic villi sample from a 37-year-old woman at the 10th week of pregnancy. She had two previous children suffering AT and two induced abortions. At molecular analysis her affected children were compound heterozygotes for mutations 7792C-->T in exon 55 (from the mother) and 8283delTC in exon 59 (from the father). The prenatal diagnosis was performed by two different operators in double-blind form. Mutation 7792C-->T was studied by restriction enzyme analysis using TaqI. Mutation 8283delTC was screened by heteroduplex analysis. The fetus was heterozygous for the mutation 7792C-->T (confirmed by sequencing). In order to verify the possible contamination by maternal DNA, polymorphic loci HLA-DRB1 and HLA-DQA1, together with microsatellite markers D6S259, D11S2000, D11S29, D11S1778 and D11S2179, were examined. All these loci were informative, showing that the fetus received only one allele from each parent. The heterozygosity for ATM mutation 7792C-->T was confirmed by molecular studies after the birth of a healthy male baby.


Assuntos
Ataxia Telangiectasia/diagnóstico , Ataxia Telangiectasia/genética , Análise Mutacional de DNA , Heterozigoto , Diagnóstico Pré-Natal/métodos , Adulto , Amostra da Vilosidade Coriônica , Feminino , Genótipo , Humanos , Recém-Nascido , Masculino , Mutação Puntual , Polimorfismo Genético , Gravidez
11.
Am J Hum Genet ; 62(3): 551-61, 1998 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-9497252

RESUMO

Ataxia-telangiectasia (A-T) is an autosomal recessive disorder characterized by cerebellar degeneration, immunodeficiency, chromosomal instability, radiosensitivity, and cancer predisposition. A-T cells are sensitive to ionizing radiation and radiomimetic chemicals and fail to activate cell-cycle checkpoints after treatment with these agents. The responsible gene, ATM, encodes a large protein kinase with a phosphatidylinositol 3-kinase-like domain. The typical A-T phenotype is caused, in most cases, by null ATM alleles that truncate or severely destabilize the ATM protein. Rare patients with milder manifestations of the clinical or cellular characteristics of the disease have been reported and have been designated "A-T variants." A special variant form of A-T is A-TFresno, which combines a typical A-T phenotype with microcephaly and mental retardation. The possible association of these syndromes with ATM is both important for understanding their molecular basis and essential for counseling and diagnostic purposes. We quantified ATM-protein levels in six A-T variants, and we searched their ATM genes for mutations. Cell lines from these patients exhibited considerable variability in radiosensitivity while showing the typical radioresistant DNA synthesis of A-T cells. Unlike classical A-T patients, these patients exhibited 1%-17% of the normal level of ATM. The underlying ATM genotypes were either homozygous for mutations expected to produce mild phenotypes or compound heterozygotes for a mild and a severe mutation. An A-TFresno cell line was found devoid of the ATM protein and homozygous for a severe ATM mutation. We conclude that certain "A-T variant" phenotypes represent ATM mutations, including some of those without telangiectasia. Our findings extend the range of phenotypes associated with ATM mutations.


Assuntos
Ataxia Telangiectasia/genética , Proteínas Serina-Treonina Quinases , Adulto , Proteínas Mutadas de Ataxia Telangiectasia , Western Blotting , Proteínas de Ciclo Celular , Linhagem Celular , Criança , Proteínas de Ligação a DNA , Feminino , Genótipo , Humanos , Linfócitos/efeitos da radiação , Masculino , Mutação , Linhagem , Fenótipo , Proteínas/genética , Tolerância a Radiação , Proteínas Supressoras de Tumor
12.
Acta Genet Med Gemellol (Roma) ; 39(4): 473-7, 1990.
Artigo em Inglês | MEDLINE | ID: mdl-1983279

RESUMO

The use of DNA restriction fragment length polymorphisms (RFLP) to analyze variable number of tandem repeat (VNTR) sequences dispersed in the human genome, has become a powerful tool for the study of population genetics due to the very substantial polymorphism involved. Because the markers usually employed for twin zygosity determination (such as sex combination, placentation, HLA typing, blood group antigens, etc) may not be uniformly informative, we propose the use of synthetic olygonucleotides, representing VNTR "core" sequences, for the determination of zygosity in twins.


Assuntos
Polimorfismo de Fragmento de Restrição , Sequências Repetitivas de Ácido Nucleico , Gêmeos/genética , Sequência de Bases , Antígenos de Grupos Sanguíneos/genética , Marcadores Genéticos , Antígenos HLA/genética , Humanos , Dados de Sequência Molecular , Sondas de Oligonucleotídeos , Gêmeos Dizigóticos , Gêmeos Monozigóticos
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