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1.
Am J Hum Genet ; 104(2): 213-228, 2019 02 07.
Artigo em Inglês | MEDLINE | ID: mdl-30639323

RESUMO

Primary defects in lung branching morphogenesis, resulting in neonatal lethal pulmonary hypoplasias, are incompletely understood. To elucidate the pathogenetics of human lung development, we studied a unique collection of samples obtained from deceased individuals with clinically and histopathologically diagnosed interstitial neonatal lung disorders: acinar dysplasia (n = 14), congenital alveolar dysplasia (n = 2), and other lethal lung hypoplasias (n = 10). We identified rare heterozygous copy-number variant deletions or single-nucleotide variants (SNVs) involving TBX4 (n = 8 and n = 2, respectively) or FGF10 (n = 2 and n = 2, respectively) in 16/26 (61%) individuals. In addition to TBX4, the overlapping ∼2 Mb recurrent and nonrecurrent deletions at 17q23.1q23.2 identified in seven individuals with lung hypoplasia also remove a lung-specific enhancer region. Individuals with coding variants involving either TBX4 or FGF10 also harbored at least one non-coding SNV in the predicted lung-specific enhancer region, which was absent in 13 control individuals with the overlapping deletions but without any structural lung anomalies. The occurrence of rare coding variants involving TBX4 or FGF10 with the putative hypomorphic non-coding SNVs implies a complex compound inheritance of these pulmonary hypoplasias. Moreover, they support the importance of TBX4-FGF10-FGFR2 epithelial-mesenchymal signaling in human lung organogenesis and help to explain the histopathological continuum observed in these rare lethal developmental disorders of the lung.

2.
Eur J Hum Genet ; 2018 Oct 29.
Artigo em Inglês | MEDLINE | ID: mdl-30374058

RESUMO

The organization of mammalian genomes into sub-megabase sized Topologically Associated Domains (TADs) has recently been revealed by techniques derived from Chromosome Conformation Capture (3 C), such as High Chromosome Contact map (Hi-C). Disruption of this organization by structural variations can lead to ectopic interactions between enhancers and promoters, and to alteration of genes expression patterns. This mechanism has already been described as the main pathophysiological mechanism in several syndromes with congenital malformations. We describe here the case of a fetus with a severe multiple congenital anomalies syndrome, including extensive polydactyly of the four limbs. This fetus carries a de novo deletion next to the IHH gene, encompassing a TAD boundary. Such an IHH TAD boundary deletion has already been described in the Dbf mouse model, which shows a quite similar, but less severe phenotype. We hypothesize that the deletion harbored by this fetus results in the same pathophysiological mechanisms as those of the Dbf model. The description of this case expands the spectrum of the disruption of chromatin architecture of WNT6/IHH/EPHA4/PAX3 locus, and could help to understand the mechanisms of chromatin interactions at this locus.

3.
Afr Health Sci ; 18(1): 147-156, 2018 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-29977268

RESUMO

Background: Truncus arteriosus communis (TAC) is a congenital heart defect in which the physiologic arterial common trunk was not divided into aorta and pulmonary artery trunk. Objectives: In this paper, we report on three observed cases from which we looked for (in conjunction with literature review) the different causes of TAC many of which have genetic origins. Methods: We collected three clinical files of fetuses having a TAC. Two of them were examinated after a medical termination of pregnancy motivated by severe cardiopathy. The malformation had been diagnosed based on different techniques: echocardiography, skeletal radiography, arteriography, fetal autopsy, karyotype and fluorescence in situ hybridization (FISH). Results: Imaging and fetopathological examination revealed the presence of TAC type 3 and 4 in the Van Praaghs classification. FISH analysis showed a 22q11.2 deletion in one fetus in favour of Digeorge syndrome. The karyotype analysis performed in two cases was normal. Conclusion: Truncus arteriosus is a rare pathology caused by numerous etiologies from which many of them have genetic origin. This malformation can be diagnosed early during prenatal period. Postmortem fetopathological examination allows a better diagnosis approach and eventually a genetic counseling in recurrent cases such as case of consanguinity.


Assuntos
Ecocardiografia/métodos , Feto/diagnóstico por imagem , Cardiopatias Congênitas/diagnóstico por imagem , Cardiopatias Congênitas/genética , Hibridização in Situ Fluorescente/métodos , Ultrassonografia Pré-Natal/métodos , Angiografia , Autopsia , Pré-Escolar , Feminino , Humanos , Lactente , Recém-Nascido , Cariotipagem , Masculino
4.
Eur J Med Genet ; 60(11): 605-609, 2017 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-28811189

RESUMO

Koolen-de Vries syndrome (MIM#610443) is a rare microdeletion syndrome involving the 17q21.31 region, which was first described by Koolen in 2006. Clinical and behavioral characteristics have been extensively reported from more than 100 postnatal cases including infants, children and young adults. The syndrome is highly clinically heterogeneous, but the main features associate characteristic cranio-facial dysmorphism, heart defects, limb, skeletal, genito-urinary anomalies, along with intellectual disability with early childhood epilepsy and behavioral disturbances. Central nervous system malformations usually consist in hydrocephalus and thin corpus callosum. We report herein an early fetal case with an apparently isolated abnormal corpus callosum diagnosed by ultrasonography, for which a medical termination of the pregnancy was achieved at 22 weeks of gestation. Postmortem examination displayed facial dysmorphism consisting of hypertelorism, short philtrum and flat and broad nose, cleft palate and left duplex ureter. Neuropathological examination revealed a mega corpus callosum that has never been reported so far in this syndrome. Array-CGH performed on thymic DNA tissue revealed a 17q21.31 microdeletion, which allowed for the confirmation of early occurring Koolen-de Vries syndrome.


Assuntos
Anormalidades Múltiplas/genética , Deficiência Intelectual/genética , Fenótipo , Anormalidades Múltiplas/diagnóstico por imagem , Anormalidades Múltiplas/patologia , Adulto , Deleção Cromossômica , Cromossomos Humanos Par 17/genética , Corpo Caloso/diagnóstico por imagem , Corpo Caloso/patologia , Feminino , Humanos , Deficiência Intelectual/diagnóstico por imagem , Deficiência Intelectual/patologia , Gravidez , Ultrassonografia Pré-Natal
5.
PLoS Genet ; 13(3): e1006683, 2017 03.
Artigo em Inglês | MEDLINE | ID: mdl-28346496

RESUMO

Schinzel-Giedion syndrome (SGS) is a rare developmental disorder characterized by multiple malformations, severe neurological alterations and increased risk of malignancy. SGS is caused by de novo germline mutations clustering to a 12bp hotspot in exon 4 of SETBP1. Mutations in this hotspot disrupt a degron, a signal for the regulation of protein degradation, and lead to the accumulation of SETBP1 protein. Overlapping SETBP1 hotspot mutations have been observed recurrently as somatic events in leukemia. We collected clinical information of 47 SGS patients (including 26 novel cases) with germline SETBP1 mutations and of four individuals with a milder phenotype caused by de novo germline mutations adjacent to the SETBP1 hotspot. Different mutations within and around the SETBP1 hotspot have varying effects on SETBP1 stability and protein levels in vitro and in in silico modeling. Substitutions in SETBP1 residue I871 result in a weak increase in protein levels and mutations affecting this residue are significantly more frequent in SGS than in leukemia. On the other hand, substitutions in residue D868 lead to the largest increase in protein levels. Individuals with germline mutations affecting D868 have enhanced cell proliferation in vitro and higher incidence of cancer compared to patients with other germline SETBP1 mutations. Our findings substantiate that, despite their overlap, somatic SETBP1 mutations driving malignancy are more disruptive to the degron than germline SETBP1 mutations causing SGS. Additionally, this suggests that the functional threshold for the development of cancer driven by the disruption of the SETBP1 degron is higher than for the alteration in prenatal development in SGS. Drawing on previous studies of somatic SETBP1 mutations in leukemia, our results reveal a genotype-phenotype correlation in germline SETBP1 mutations spanning a molecular, cellular and clinical phenotype.


Assuntos
Anormalidades Múltiplas/genética , Proteínas de Transporte/genética , Anormalidades Craniofaciais/genética , Predisposição Genética para Doença/genética , Deformidades Congênitas da Mão/genética , Neoplasias Hematológicas/genética , Deficiência Intelectual/genética , Mutação , Unhas Malformadas/genética , Proteínas Nucleares/genética , Anormalidades Múltiplas/metabolismo , Anormalidades Múltiplas/patologia , Western Blotting , Proteínas de Transporte/metabolismo , Linhagem Celular , Proliferação de Células/genética , Transformação Celular Neoplásica/genética , Criança , Pré-Escolar , Anormalidades Craniofaciais/metabolismo , Anormalidades Craniofaciais/patologia , Feminino , Perfilação da Expressão Gênica , Estudos de Associação Genética , Mutação em Linhagem Germinativa , Células HEK293 , Deformidades Congênitas da Mão/metabolismo , Deformidades Congênitas da Mão/patologia , Neoplasias Hematológicas/metabolismo , Neoplasias Hematológicas/patologia , Humanos , Lactente , Recém-Nascido , Deficiência Intelectual/metabolismo , Deficiência Intelectual/patologia , Masculino , Unhas Malformadas/metabolismo , Unhas Malformadas/patologia , Proteínas Nucleares/metabolismo , Fenótipo
6.
Prenat Diagn ; 36(13): 1270-1275, 2016 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-27859469

RESUMO

OBJECTIVE: Fraser syndrome (FS) is a rare malformation recessive disorder. Major criteria are cryptophtalmos, syndactyly, respiratory, genital and urinary tract anomalies. Few prenatal presentations have been reported. METHOD: We analyzed the prenatal and postnatal fetal phenotype in 38 cases of FS, including 25 pregnancy termination cases, 8 intra-uterine death cases and 4 cases that died after birth. RESULTS: Including both prenatal and postnatal fetal phenotypic evaluation, all cases presented dysmorphic features with nose and ear dysplasia. Renal anomalies and syndactyly were present in 37/38 cases, cryptophtalmos in 36/38, airways anomalies in 30/37 and genital anomalies in 30/35 cases. Anomalies of the abdominal wall such as low set umbilicus and omphalocele were found in 31 cases. Among the 26 cases for which ultrasound data were available, detectable anomalies included oligohydramnios (22), ascites/hydrops (9), renal anomalies (20), evidence for high airways obstruction (11), ophthalmologic anomalies (4), ear dysplasia (2) and syndactyly (2). CONCLUSION: This study shows that the postnatal phenotype of FS is very specific, whereas oligohydramnios hampers the prenatal recognition of the cardinal FS diagnosis criteria. Association of oligohydramnios, kidney agenesis and CHAOS should lead to consider this diagnosis. © 2016 John Wiley & Sons, Ltd.


Assuntos
Anormalidades Múltiplas/diagnóstico , Anormalidades Múltiplas/embriologia , Síndrome de Fraser/diagnóstico , Síndrome de Fraser/embriologia , Diagnóstico Pré-Natal/métodos , Obstrução das Vias Respiratórias/diagnóstico por imagem , Obstrução das Vias Respiratórias/embriologia , Anormalidades Congênitas/diagnóstico por imagem , Anormalidades Congênitas/embriologia , Anormalidades Craniofaciais/diagnóstico , Anormalidades Craniofaciais/embriologia , Orelha/anormalidades , Orelha/diagnóstico por imagem , Orelha/embriologia , Anormalidades do Olho/diagnóstico por imagem , Anormalidades do Olho/embriologia , Feminino , Síndrome de Fraser/diagnóstico por imagem , Idade Gestacional , Humanos , Hidropisia Fetal/diagnóstico por imagem , Recém-Nascido , Rim/anormalidades , Rim/diagnóstico por imagem , Rim/embriologia , Oligo-Hidrâmnio/diagnóstico por imagem , Fenótipo , Gravidez , Sindactilia/diagnóstico por imagem , Ultrassonografia Pré-Natal , Anormalidades Urogenitais/diagnóstico
7.
Urology ; 89: 132-3, 2016 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-26657689

RESUMO

Prenatal testicular torsion is a very rare morbid entity, described in the literature to occur when the testicle is intrascrotal, around the 34th week of gestation. Here we report a case of early testicular necrosis. This male fetus was the product of a medical abortion at 27 weeks. During evisceration, a left testicular nubbin free in the peritoneal cavity was found. Histologically, it was extensively necrotic. Because of the location, the size, and the histological features of this necrotic testicle, we conclude that it was the result of torsion of the pedicle that occurred around the 20th week of pregnancy.


Assuntos
Torção do Cordão Espermático/embriologia , Feminino , Idade Gestacional , Humanos , Masculino , Gravidez , Terceiro Trimestre da Gravidez
8.
Eur J Hum Genet ; 23(1): 92-102, 2015 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-24736735

RESUMO

The phenotypic spectrum of GLI3 mutations includes autosomal dominant Greig cephalopolysyndactyly syndrome (GCPS) and Pallister-Hall syndrome (PHS). PHS was first described as a lethal condition associating hypothalamic hamartoma, postaxial or central polydactyly, anal atresia and bifid epiglottis. Typical GCPS combines polysyndactyly of hands and feet and craniofacial features. Genotype-phenotype correlations have been found both for the location and the nature of GLI3 mutations, highlighting the bifunctional nature of GLI3 during development. Here we report on the molecular and clinical study of 76 cases from 55 families with either a GLI3 mutation (49 GCPS and 21 PHS), or a large deletion encompassing the GLI3 gene (6 GCPS cases). Most of mutations are novel and consistent with the previously reported genotype-phenotype correlation. Our results also show a correlation between the location of the mutation and abnormal corpus callosum observed in some patients with GCPS. Fetal PHS observations emphasize on the possible lethality of GLI3 mutations and extend the phenotypic spectrum of malformations such as agnathia and reductional limbs defects. GLI3 expression studied by in situ hybridization during human development confirms its early expression in target tissues.


Assuntos
Estudos de Associação Genética , Fatores de Transcrição Kruppel-Like/genética , Mutação , Proteínas do Tecido Nervoso/genética , Anormalidades Múltiplas/diagnóstico , Anormalidades Múltiplas/genética , Acrocefalossindactilia/diagnóstico , Acrocefalossindactilia/genética , Estudos de Coortes , Análise Mutacional de DNA , Família , Expressão Gênica , Rearranjo Gênico , Haploinsuficiência , Humanos , Hibridização in Situ Fluorescente , Fenótipo , Proteína Gli3 com Dedos de Zinco
9.
Ann Pathol ; 34(2): 119-23, 2014 Apr.
Artigo em Francês | MEDLINE | ID: mdl-24703021

RESUMO

Of all the gestational trophoblastic tumors, the gestational choriocarcinomas have the worst prognosis and the most uncommon. We report a case diagnosed on a full-term placenta, discovered incidentally. The patient, gravida 2, para 1, delivered a hypotrophic infant at 38 weeks gestation. The placenta was examined in the laboratory to perform anatomo-pathological examination in order to explain the growth retardation. This study revealed the presence of an intraplacental choriocarcinoma. Disease staging was negative for both mother and child, and beta-HCG levels remained at zero. These two factors are rather good prognosis for choriocarcinoma. With this observation, we highlight the added-value of placental examination, which seems essential for any fetal pathology, pathological pregnancy and intrapartum complications. Anatomo-pathological examination must be meticulous and systematized in order to not overlook an intraplacental tumor.


Assuntos
Coriocarcinoma/patologia , Doenças Placentárias/patologia , Adulto , Feminino , Humanos , Achados Incidentais , Gravidez , Nascimento a Termo
10.
Pediatr Dev Pathol ; 17(2): 102-6, 2014 Mar-Apr.
Artigo em Inglês | MEDLINE | ID: mdl-24575782

RESUMO

Feto-maternal hemorrhage (FMH) is the cause of late fetal death in 1.6%-11% of cases. In spite of this high frequency, its pathological features have received little attention. The definitive diagnosis of lethal FMH requires confirmation of sufficient fetal blood volume loss. This is determined by tests such as the Kleihauer-Betke test, which may not have been obtained or not have been available before the autopsy. The pathologist may offer a tentative diagnosis of FMH from the autopsy findings. The objective of this study was to better characterize the placental and fetal autopsy findings in lethal FMH. This was a retrospective study of 17 cases of FMH proven by a positive Kleihauer-Betke test. The cases were selected from the autopsy files of the Department of Pathology, Centre Hospitalier Universitaire de Bordeaux. The pathological reports as well as the placental and fetal photographs and the microscopic slides of each case were systematically reviewed. The fetal autopsy findings in FMH are characterized by a eutrophic pale macerated fetus, low liver weight, absent intrathoracic petechiae, increased extramedullary hematopoiesis in the liver and kidney, and increased circulating nucleated red blood cells. The placenta shows an increased frequency of intervillous thrombi. Although nonpathognomonic, some of the pathological features are strongly suggestive of FMH. When the latter is present, a Kleihauer-Betke test should be performed, even some days after the delivery.


Assuntos
Morte Fetal/etiologia , Feto/patologia , Hemorragia/patologia , Placenta/patologia , Adulto , Autopsia/métodos , Feminino , Morte Fetal/metabolismo , Idade Gestacional , Hemorragia/diagnóstico , Humanos , Placenta/irrigação sanguínea , Gravidez
11.
Ann Pathol ; 33(4): 230-6, 2013 Aug.
Artigo em Francês | MEDLINE | ID: mdl-23954115

RESUMO

The indications of the pathological examination of the placenta are mainly represented by uteroplacental vascular deficiency. The clinical context is often evocative, but it can sometimes be solely an intra-uterine growth retardation or an unexplained in utero fetal death. So, the pathological lesions of this uteroplacental vascular deficiency must be well-known to be correctly interpreted, for none of these lesions is truly specific. The pathological diagnosis is based on a group of macroscopic and microscopic arguments. Various physiopathological mechanisms, often imperfectly known, can be at the origin of an uteroplacental vascular insufficiency, but in the current position, the pathological examination does not allow etiopathogenic orientation. The development of the trophoblastic biopsies gives us access to a new material which, in parallel with the cytogenetic analysis, often allows us, in front of an unexplained intra-uterine growth retardation, to direct the diagnosis towards uteroplacental vascular insufficiency. The histological analysis of the chorionic villous sampling taken precociously during pathological pregnancies is thus a major diagnostic contribution. But especially, this analysis gives access to new information which, in the near future, will enable us to better define the pathological evolution of the lesions of hypoxic chorionic villous and to contribute to a better knowledge of this pathology which, under many aspects, still conceals many mysteries.


Assuntos
Doenças Placentárias/patologia , Placenta/patologia , Circulação Placentária , Útero/patologia , Vilosidades Coriônicas/química , Vilosidades Coriônicas/patologia , Amostra da Vilosidade Coriônica , Cistos/patologia , Feminino , Morte Fetal/patologia , Hipóxia Fetal/etiologia , Fibrina/análise , Idade Gestacional , Humanos , Infarto/patologia , Necrose , Tamanho do Órgão , Placenta/irrigação sanguínea , Gravidez , Complicações na Gravidez/fisiopatologia , Trofoblastos/patologia , Útero/irrigação sanguínea
12.
Ann Pathol ; 32(3): 217-9, 2012 Jun.
Artigo em Francês | MEDLINE | ID: mdl-22748341

RESUMO

Tamponade is a rare but particularly serious complication of central venous catheters in the newborn. Tamponade can be due to the endocardic aggression caused by the continuous flow of a hyperosmotic solution or by a mechanical injury that can result in perforation of the atrial wall. The risk of tamponade is present whatever is the position of the tip of the catheter, although it has been shown that this risk is increased when this tip is in the right auricle. The originality of our observation is the discovery at the post-mortem examination of an anterior interventricular vein thrombosis, without any lesion of the atrial wall. In the event of the diagnosis of tamponade in living newborn, this etiology must be required because of its therapeutic implications.


Assuntos
Tamponamento Cardíaco/etiologia , Tamponamento Cardíaco/patologia , Cateterismo Venoso Central/efeitos adversos , Doenças em Gêmeos/etiologia , Doenças em Gêmeos/patologia , Trombose Venosa/etiologia , Trombose Venosa/patologia , Evolução Fatal , Feminino , Ventrículos do Coração , Humanos , Recém-Nascido
13.
Ann Surg Oncol ; 18(5): 1372-9, 2011 May.
Artigo em Inglês | MEDLINE | ID: mdl-21108045

RESUMO

BACKGROUND: The objective of this retrospective study was to identify prognostic, diagnostic, and therapeutic disparities between younger (≤ 40 years) and older (> 40 years) women with ductal carcinoma in situ (DCIS) of the breast. METHODS: From 1971 to 2001, all patients treated for DCIS at Institut Bergonié were included in our analyses. Follow-up data was collected over 10 years. We used univariate and multivariate analyses to investigate patient-, disease-, and treatment-related factors predictive of diagnostic, histological, therapeutic, and prognostic DCIS criteria. RESULTS: A total of 812 patients were eligible including 731 women aged >40 years and 81 women ≤40 years. Younger women with DCIS were more likely to receive a mastectomy and less likely to receive radiotherapy. Young age and initial surgical treatment (lumpectomy and especially nonfree margins) were revealed as predictive of recurrence in multivariate analyses. CONCLUSIONS: Young age represents a recurrence risk independent of histological and clinical characteristics of the tumor. Initial treatment, especially for nonfree margins, is also a predictive factor. Appropriate initial surgery with particularly wide margins appears essential for the treatment of young women with DCIS.


Assuntos
Neoplasias da Mama/mortalidade , Carcinoma Ductal de Mama/mortalidade , Carcinoma Intraductal não Infiltrante/mortalidade , Carcinoma Lobular/mortalidade , Recidiva Local de Neoplasia/mortalidade , Adulto , Fatores Etários , Idoso , Idoso de 80 Anos ou mais , Neoplasias da Mama/diagnóstico , Neoplasias da Mama/cirurgia , Carcinoma Ductal de Mama/diagnóstico , Carcinoma Ductal de Mama/cirurgia , Carcinoma Intraductal não Infiltrante/diagnóstico , Carcinoma Intraductal não Infiltrante/cirurgia , Carcinoma Lobular/diagnóstico , Carcinoma Lobular/cirurgia , Feminino , Seguimentos , Humanos , Pessoa de Meia-Idade , Invasividade Neoplásica , Recidiva Local de Neoplasia/diagnóstico , Recidiva Local de Neoplasia/cirurgia , Prognóstico , Estudos Retrospectivos , Fatores de Risco , Taxa de Sobrevida
14.
Ann Pathol ; 29(5): 424-7, 2009 Oct.
Artigo em Francês | MEDLINE | ID: mdl-20004848

RESUMO

A case of maze-like angiomatoid anomaly in villi obtained by chorionic villous sampling (CVS) is described. This feature is pathognomonic of partial mole (triploid syndrome) and it was later confirmed by chromosomal analysis. Maze-like angiomatoid anomaly was previously described on specimen submitted after spontaneous or induced abortions, but it was never reported on CVS. This report emphasized that microscopic investigation of CVS cannot be conclusive for cytogenetic anomaly in almost all cases excepted for partial mole where diagnosis criteria are usually characteristic.


Assuntos
Amostra da Vilosidade Coriônica , Mola Hidatiforme/patologia , Neoplasias Uterinas/patologia , Adulto , Feminino , Humanos , Gravidez
15.
Biomacromolecules ; 7(4): 1147-55, 2006 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-16602732

RESUMO

In a previous paper we reported that beta-D-glucans isolated from Saccharomyces cerevisiae could adsorb zearalenone, reduce its bioavailability in the digestive tract, and protect animals against its adverse effects. We have now investigated, in vitro, the kinetics of the interaction between other mycotoxins and beta-D-glucans from several sources at three pH values found along the digestive tract (3.0, 6.0, and 8.0). Acid and neutral conditions gave the highest affinity rates for aflatoxins B1 > deoxynivalenol > ochratoxin A and involved both the (1 --> 3)-beta-D-glucans and the (1 --> 6)-beta-D-glucans. Alkaline conditions, owing to their destructuring action on glucans, were favorable only for the adsorption of patulin. Using molecular mechanics, we found that hydroxyl, ketone, and lactone groups are involved in the formation of both hydrogen bonds and van der Waals interactions between aflatoxins B1, deoxynivalenol and patulin, and beta-D-glucans. Differences in the binding capacity of the mycotoxins are due to their specific physical and chemical characteristics.


Assuntos
Micotoxinas/química , Conformação Proteica , beta-Glucanas/química , Adsorção , Sítios de Ligação , Parede Celular/química , Ligações de Hidrogênio , Concentração de Íons de Hidrogênio , Técnicas In Vitro , Cinética , Modelos Moleculares , Estrutura Molecular , Saccharomyces cerevisiae/química , Dióxido de Silício/química , Propriedades de Superfície
16.
Biomacromolecules ; 5(6): 2176-85, 2004 Nov-Dec.
Artigo em Inglês | MEDLINE | ID: mdl-15530031

RESUMO

The beta-D-glucans from the cell wall of Saccharomyces cerevisiae have shown in vitro affinity for zearalenone. For this reason, their utilization as dietary adsorbent, to reduce the bioavailability of zearalenone, is of practical interest. Our study used powerful devices to elucidate the spatial conformation and molecular sites of interaction between ZEN and beta-D-glucans. In this respect, 1H NMR spectroscopy implicated the hydroxyl groups of the phenol moiety of zearalenone in the complexation by laminarin, a pure beta-(1,3)-D-glucan. X-ray diffraction determined that laminarin displays the conformation of a single-helix with six beta-D-glucopyranose residues per turn. At this stage, molecular modeling was useful to locate the interaction sites and to propose highly probable complexes of zearalenone with laminarin fragment. Interestingly, the beta-(1,3)-D-glucan chain favors a very stable intra-helical association with zearalenone, nicely stabilized by beta-(1,6)-D-glucans side chains. Both hydrogen bonds and van der Waals interactions were precisely identified in the complex and could thus be proposed as driving interactions to monitor the association between the two molecules.


Assuntos
Zearalenona/química , beta-Glucanas/química , Dissacarídeos/química , Glucanos/química , Ligações de Hidrogênio , Espectroscopia de Ressonância Magnética , Modelos Químicos , Modelos Moleculares , Polímeros/química , Conformação Proteica , Saccharomyces cerevisiae/metabolismo , Software , Difração de Raios X
17.
J Biol Chem ; 279(1): 726-34, 2004 Jan 02.
Artigo em Inglês | MEDLINE | ID: mdl-14570882

RESUMO

Amylosucrase from Neisseria polysaccharea is a remarkable transglucosidase from family 13 of the glycoside-hydrolases that synthesizes an insoluble amylose-like polymer from sucrose in the absence of any primer. Amylosucrase shares strong structural similarities with alpha-amylases. Exactly how this enzyme catalyzes the formation of alpha-1,4-glucan and which structural features are involved in this unique functionality existing in family 13 are important questions still not fully answered. Here, we provide evidence that amylosucrase initializes polymer formation by releasing, through sucrose hydrolysis, a glucose molecule that is subsequently used as the first acceptor molecule. Maltooligosaccharides of increasing size were produced and successively elongated at their nonreducing ends until they reached a critical size and concentration, causing precipitation. The ability of amylosucrase to bind and to elongate maltooligosaccharides is notably due to the presence of key residues at the OB1 acceptor binding site that contribute strongly to the guidance (Arg415, subsite +4) and the correct positioning (Asp394 and Arg446, subsite +1) of acceptor molecules. On the other hand, Arg226 (subsites +2/+3) limits the binding of maltooligosaccharides, resulting in the accumulation of small products (G to G3) in the medium. A remarkable mutant (R226A), activated by the products it forms, was generated. It yields twice as much insoluble glucan as the wild-type enzyme and leads to the production of lower quantities of by-products.


Assuntos
Glucosiltransferases/química , Glucosiltransferases/metabolismo , Neisseria/enzimologia , Oligossacarídeos/metabolismo , Sequência de Aminoácidos , Substituição de Aminoácidos , Sítios de Ligação , Sequência Conservada , Glucosiltransferases/isolamento & purificação , Modelos Moleculares , Mutagênese Sítio-Dirigida , Conformação Proteica , Proteínas Recombinantes/química , Proteínas Recombinantes/metabolismo , Alinhamento de Sequência , Homologia de Sequência de Aminoácidos , Sacarose/metabolismo
18.
J Biol Chem ; 279(11): 10093-102, 2004 Mar 12.
Artigo em Inglês | MEDLINE | ID: mdl-14660599

RESUMO

The role in activity of outer regions in the substrate binding cleft in alpha-amylases is illustrated by mutational analysis of Tyr(105) and Thr(212) localized at subsites -6 and +4 (substrate cleavage occurs between subsites -1 and +1) in barley alpha-amylase 1 (AMY1). Tyr(105) is conserved in plant alpha-amylases whereas Thr(212) varies in these and related enzymes. Compared with wild-type AMY1, the subsite -6 mutant Y105A has 140, 15, and <1% activity (k(cat)/K(m)) on starch, amylose DP17, and 2-chloro-4-nitrophenyl beta-d-maltoheptaoside, whereas T212Y at subsite +4 has 32, 370, and 90% activity, respectively. Thus engineering of aromatic stacking interactions at the ends of the 10-subsite long binding cleft affects activity very differently, dependent on the substrate. Y105A dominates in dual subsite -6/+4 [Y105A/T212(Y/W)]AMY1 mutants having almost retained and low activity on starch and oligosaccharides, respectively. Bond cleavage analysis of oligosaccharide degradation by wild-type and mutant AMY1 supports that Tyr(105) is critical for binding at subsite -6. Substrate binding is improved by T212(Y/W) introduced at subsite +4 and the [Y105A/T212(Y/W)]AMY1 double mutants synergistically enhanced productive binding of the substrate aglycone. The enzymatic properties of the series of AMY1 mutants suggest that longer substrates adopt several binding modes. This is in excellent agreement with computed distinct multiple docking solutions observed for maltododecaose at outer binding areas of AMY1 beyond subsites -3 and +3.


Assuntos
Hordeum/enzimologia , Oligossacarídeos/química , Treonina/química , Tirosina/química , alfa-Amilases/química , Sequência de Aminoácidos , Sítios de Ligação , Domínio Catalítico , Eletroforese em Gel de Poliacrilamida , Escherichia coli/metabolismo , Ligações de Hidrogênio , Hidrólise , Cinética , Modelos Moleculares , Dados de Sequência Molecular , Mutagênese Sítio-Dirigida , Mutação , Peptídeos/química , Pichia/metabolismo , Plasmídeos/metabolismo , Ligação Proteica , Proteínas Recombinantes/química , Especificidade por Substrato
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