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1.
Article in Chinese | WPRIM | ID: wpr-921983

ABSTRACT

OBJECTIVE@#To investigate the clinical features and SLC35A2 variant of a case of congenital disorder of glycosylation type IIm (CDG-IIm), and to identify the possible causes of the disease.@*METHODS@#Trio-whole exome sequencing (WES) was used to analyze the gene variant of the children and their parents. The suspicious gene variants were screened for Sanger verification and the bioinformatics prediction was used to analyze the hazard of variant.@*RESULTS@#The clinical manifestations of the child were epilepsy, global growth retardation, nystagmus, myocarditis and other symptoms. MRI showed brain dysplasia such as wide frontal temporal sulcus and subarachnoid space on both sides. Echocardiography showed left ventricular wall thickening and patent foramen ovale. According to the results of gene detection, there was a heterozygous missense variant c.335C>A (p.Thr112Lys) in SLC35A2 gene. The parents were wild-type at this locus, which was a de novo variant. At the same time, there was no report of this variant in the relevant literature, which was a novel variant in SLC35A2 gene. According to the genetic variant guidelines of American College of Medical Genetics and Genomics, SLC35A2 gene c.335C>A (p.Thr112Lys) variant was predicted to be likely pathogenic (PS2+PM2+PP3).@*CONCLUSION@#The variant of SLC35A2 gene c.335C>A(p.Thr112Lys) may be the cause of the disease in the child.


Subject(s)
Child , Congenital Disorders of Glycosylation/genetics , Glycosylation , Humans , Magnetic Resonance Imaging , Monosaccharide Transport Proteins/genetics , Whole Exome Sequencing
2.
Chinese Journal of Biotechnology ; (12): 4036-4046, 2021.
Article in Chinese | WPRIM | ID: wpr-921484

ABSTRACT

N-glycosylation modification, one of the most common protein post-translational modifications, occurs in heat shock protein gp96. The purpose of this study is to investigate the effect of N-glycosylation modification on immunologic function of the recombinant gp96 using the mutant gp96 in N-glycosylation sites. Firstly, wild-type and mutant gp96 proteins were expressed by insect expression system and their glycosylation levels were detected. To determine the effect of N-glycosylation on gp96 antigen presentation function, the IFN-γ+ CD8+ T cells in gp96-immunized mice and secretion level of IFN-γ were examined by flow cytometry and ELISA. The ATPase activity of gp96 was further detected by the ATPase kit. Finally, the effect of N-glycosylation on adjuvant function of gp96 for influenza vaccine was investigated in immunized mice. It was found that total sugar content of mutant recombinant gp96 was reduced by 27.8%. Compared to the wild type recombinant gp96, mutations in N-glycosylation sites resulted in decreased antigen presentation ability and ATPase activity of gp96. Furthermore, influenza vaccine-specific T cell levels induced by mutant gp96 as adjuvant were dramatically reduced compared to those by wild type recombinant gp96. These results demonstrate that N-glycosylation modification is involved in regulation of ATPase activity and antigen presentation function of gp96, thereby affecting its adjuvant function. The results provide the technical bases for development of gp96- adjuvanted vaccines.


Subject(s)
Adjuvants, Immunologic , Animals , CD8-Positive T-Lymphocytes/metabolism , Glycosylation , Heat-Shock Proteins , Influenza Vaccines , Mice
3.
Article in Chinese | WPRIM | ID: wpr-888017

ABSTRACT

Type 2 diabetes mellitus( T2 DM) is a common chronic metabolic disease characterized by persistent hyperglycemia and insulin resistance. In pancreatic β-cells,glucose-stimulated insulin secretion( GSIS) plays a pivotal role in maintaining the balance of blood glucose level. Previous studies have shown that geniposide,one of the active components of Gardenia jasminoides,could quickly regulate the absorption and metabolism of glucose,and affect glucose-stimulated insulin secretion in pancreatic β cells,but the specific mechanism needs to be further explored. Emerging evidence indicated that glycosylation of glucose transporter( GLUT) has played a key role in sensing cell microenvironmental changes and regulating glucose homeostasis in eucaryotic cells. In this study,we studied the effects of geniposide on the key molecules of GLUT2 glycosylation in pancreatic β cells. The results showed that geniposide could significantly up-regulate the mRNA and protein levels of Glc NAc T-Ⅳa glycosyltransferase( Gn T-Ⅳa) and galectin-9 but had no signi-ficant effect on the expression of clathrin,and geniposide could distinctively regulate the protein level of Gn T-Ⅳa in a short time( 1 h) under the conditions of low and medium glucose concentrations,but had no significant effect on the protein level of galectin-9. In addition,geniposide could also remarkably affect the protein level of glycosylated GLUT2 in a short-time treatment. The above results suggested that geniposide could quickly regulate the protein level of Gn T-Ⅳa,a key molecule of protein glycosylation in INS-1 rat pancreatic βcells and affect the glycosylation of GLUT2. These findings suggested that the regulation of geniposide on glucose absorption,metabolism and glucose-stimulated insulin secretion might be associated with its efficacy in regulating GLUT2 glycosylation and affecting its distribution on the cell membrane and cytoplasm in pancreatic β cells.


Subject(s)
Animals , Diabetes Mellitus, Type 2/metabolism , Glucose/metabolism , Glycosylation , Insulin/metabolism , Insulin-Secreting Cells/metabolism , Iridoids , Rats
4.
Article in Chinese | WPRIM | ID: wpr-878883

ABSTRACT

Nano-LC MS/MS was used to analyze trypsin digested deer-hide gelatin(DHG) samples, hydroxylation and O-glycosylation on lysine sites of DHG were comprehensive identified by using PEAKS Studio software. The sites, sorts and amounts of hydroxylation and O-glycosylation on Type Ⅰ collagen α1 chain(COL1 A1) and α2 chain(COL1 A2) of DHG were revealed. As a result, 5 284 peptides were identified from DHG samples, which were mainly from COL1 A1 and COL1 A2. Among these peptides, there were 449 peptides with hydroxylysine, 442 with galactosyl-hydroxylysine, 449 with glucosyl-galactosyl-hydroxylysine. The major modified sites of hydroxylation and O-glycosylation in DHG were shown as follow: α1-9 N and α2-5 N in N-telopeptides, α1-87, α1-174, α1-930, α2-87, α2-174, α2-933 in triple helix domain, and α1-16 C in C-telopeptides. These hydroxylation and O-glycosylation were correlated with the formation and stability of collagen molecules and collagen fibrils. It is feasible for the collagens and peptides dissolving from deer skin collagen fibrils under high temperature and pressure decocting, high temperature and pressure also might destroy inter-molecular covalent cross-linking and help those glycol-peptides formations. The present study provided ideas and strategies for the in-depth investigation on DHG chemical constituents, and showed good theoretical significance and application value.


Subject(s)
Animals , Deer/metabolism , Gelatin , Glycosylation , Hydroxylation , Lysine/metabolism , Protein Processing, Post-Translational , Tandem Mass Spectrometry
5.
Chinese Journal of Biotechnology ; (12): 112-129, 2021.
Article in Chinese | WPRIM | ID: wpr-878547

ABSTRACT

Water solubility, stability, and bioavailability, can be substantially improved after glycosylation. Glycosylation of bioactive compounds catalyzed by glycoside hydrolases (GHs) and glycosyltransferases (GTs) has become a research hotspot. Thanks to their rich sources and use of cheap glycosyl donors, GHs are advantageous in terms of scaled catalysis compared to GTs. Among GHs, sucrose phosphorylase has attracted extensive attentions in chemical engineering due to its prominent glycosylation activity as well as its acceptor promiscuity. This paper reviews the structure, catalytic characteristics, and directional redesign of sucrose phosphorylase. Meanwhile, glycosylation of diverse chemicals with sucrose phosphorylase and its coupling applications with other biocatalysts are summarized. Future research directions were also discussed based on the current research progress combined with our working experience.


Subject(s)
Glucosyltransferases/metabolism , Glycoside Hydrolases/metabolism , Glycosylation , Glycosyltransferases/genetics
6.
J. venom. anim. toxins incl. trop. dis ; 27: e20200182, 2021. tab, graf, ilus
Article in English | LILACS, VETINDEX | ID: biblio-1250254

ABSTRACT

The accessory ß1 subunits, regulating the pharmacological and biophysical properties of BK channels, always undergo post-translational modifications, especially glycosylation. To date, it remains elusive whether the glycosylation contributes to the regulation of BK channels by ß1 subunits. Methods: Herein, we combined the electrophysiological approach with molecular mutations and biochemical manipulation to investigate the function roles of N-glycosylation in ß1 subunits. Results: The results show that deglycosylation of ß1 subunits through double-site mutations (ß1 N80A/N142A or ß1 N80Q/N142Q) could significantly increase the inhibitory potency of iberiotoxin, a specific BK channel blocker. The deglycosylated channels also have a different sensitivity to martentoxin, another BK channel modulator with some remarkable effects as reported before. On the contrary to enhancing effects of martentoxin on glycosylated BK channels under the presence of cytoplasmic Ca2+, deglycosylated channels were not affected by the toxin. However, the deglycosylated channels were surprisingly inhibited by martentoxin under the absence of cytoplasmic Ca2+, while the glycosylated channels were not inhibited under this same condition. In addition, wild type BK (α+ß1) channels treated with PNGase F also showed the same trend of pharmacological results to the mutants. Similar to this modulation of glycosylation on BK channel pharmacology, the deglycosylated forms of the channels were activated at a faster speed than the glycosylated ones. However, the V1/2 and slope were not changed by the glycosylation. Conclusion: The present study reveals that glycosylation is an indispensable determinant of the modulation of ß1-subunit on BK channel pharmacology and its activation. The loss of glycosylation of ß1 subunits could lead to the dysfunction of BK channel, resulting in a pathological state.(AU)


Subject(s)
Glycosylation , Peptide-N4-(N-acetyl-beta-glucosaminyl) Asparagine Amidase , Mutation , Pharmacology
7.
Braz. arch. biol. technol ; 64: e21200301, 2021. tab, graf
Article in English | LILACS | ID: biblio-1278443

ABSTRACT

Abstract Rhamnolipid is a potent biodegradable surfactant, which frequently used in pharmaceutical and environmental industries, such as enhanced oil recovery and bioremediation. This study aims to engineer Escherichia coli for the heterologous host production of rhamnolipid, to characterize the rhamnolipid product, and to optimize the production using autoinduction medium and POME (palm oil mill effluent). The construction of genes involved in rhamnolipid biosynthesis was designed in two plasmids, pPM RHLAB (mono-rhamnolipid production plasmid) and pPM RHLABC (di-rhamnolipid production plasmid). The characterization of rhamnolipid congeners and activity using high-resolution mass spectrometry (HRMS) and critical micelle concentration (CMC). In order to estimate rhamnolipid yield, an oil spreading test was performed. HRMS and CMC result show E. coli pPM RHLAB mainly produced mono-rhamnolipid (Rha-C14:2) with 900 mg/L and 35.4 mN/m of CMC and surface tension value, whereas E. coli pPM RHLABC mainly produced di-rhamnolipid (Rha-Rha-C10) with 300 mg/L and 34.3 mN/m of CMC and surface tension value, respectively. The optimum condition to produce rhamnolipid was at 20 h cultivation time, 37 oC, and pH 7. In this condition, the maximum rhamnolipid yield of 1245.68 mg/L using autoinduction medium and 318.42 mg/L using 20% (v/v) of POME. In conclusion, the characteristics of the rhamnolipid by recombinant E. coli is very promising to be used in industries as the most economical way of producing rhamnolipid.


Subject(s)
Palm Oil , Escherichia coli , Electromagnetic Phenomena , Glycosylation
9.
Article in English | WPRIM | ID: wpr-811195

ABSTRACT

PURPOSE: Aberrant glycosylation of the histo-blood group antigens (including the angina bullosa haemorrhagica [ABH]) is often observed during malignant transformation in most types of carcinomas. Data concerning their ethnic distributions are diverse which explains why their biological characteristics have to be studied in different populations. Our aim was to analyze the expression of the histo-blood group (specifically the ABH) antigens in breast carcinoma.METHODS: The expression of the histo-blood group (specifically the ABH) antigens was studied in 109 patients with breast carcinoma using immunohistochemistry. Statistical analysis was performed using χ² and Fisher analyses.RESULTS: The loss of expression of histo-blood group (ABH) antigens in breast carcinoma was observed in 81.13% of patients with blood group O, 37.93% with blood group A, and 96.30% with blood group B. One key finding of this study was that the loss of expression of the ABH antigen was also observed in normal tissues adjacent to the tumor. The loss of expression was associated with higher tumor grade (p < 0.05). Expression of H antigen was observed in 50% of cases with loss of expression of B antigen and was associated with human epidermal growth factor receptor 2 (HER2) overexpression (p < 0.05). The loss of H antigen in patients with blood group O was associated with estrogen receptor expression (p < 0.001). Incompatible A antigen in tumor was expressed in 20.75% of patients with blood group O.CONCLUSION: Loss of the ABH antigens correlated with the Scarff-Bloom-Richardson histologic grading. H antigen was associated with HER2 overexpression in breast cancer. However, further studies are needed to determine the role of incompatible A antigen in mammary carcinogenesis.


Subject(s)
Breast Neoplasms , Breast , Carcinogenesis , Estrogens , Glycosylation , Histocompatibility , Humans , Immunohistochemistry , Population Characteristics , ErbB Receptors
10.
Article in Chinese | WPRIM | ID: wpr-878682

ABSTRACT

Proteins exert their roles in life activities via post-translational modifications(PTMs),which include phosphorylation,acetylation,ubiquitination,glycosylation,and methylation.These modifications can change the functions of proteins and play key roles in a variety of diseases.Endometriosis is a common disease in women of childbearing age,although its molecular mechanisms remain unclear.Recent studies have shown that PTMs may be involved in the pathogenesis of endometriosis.Here we review the roles of PTMs in the occurrence and development of endometriosis and the potential medical treatments.


Subject(s)
Acetylation , Endometriosis/pathology , Female , Glycosylation , Humans , Phosphorylation , Protein Processing, Post-Translational , Ubiquitination
11.
Chinese Journal of Biotechnology ; (12): 2313-2326, 2020.
Article in Chinese | WPRIM | ID: wpr-878488

ABSTRACT

Glycosylation is one of the common post-translational modifications of proteins to regulate the ability of tumor invasion, metastasis and tumor heterogeneity by interacting with glycan-binding proteins such as lectins and antibodies. Glycan microarray can be constructed by chemical synthesis, chemical-enzyme synthesis or natural glycan releasing. Glycan microarray is an essential analytical tool to discover the interaction between glycan and its binding proteins. Here we summarize the standard techniques to construct glycan microarray for the application in cancer vaccine, monoclonal antibody and diagnostic markers.


Subject(s)
Antibodies, Monoclonal , Glycosylation , Lectins/metabolism , Microarray Analysis , Neoplasms , Polysaccharides
12.
São Paulo; s.n; s.n; 2019. 94 p. graf, tab.
Thesis in Portuguese | LILACS | ID: biblio-1024757

ABSTRACT

L-asparaginase é um inibidor eficiente do crescimento tumoral, usado em sessões de quimioterapia contra a Leucemia Linfoblástica Aguda (LLA), resultando na remissão completa da doença em 90% dos pacientes tratados. A L-asparaginase II de Saccharomyces cerevisiae (ScASNaseII) tem alto potencial de superar os efeitos adversos da L-asparaginase de bactéria, porém sua produção endógena resulta em uma proteína hipermanosilada e, consequentemente, imunogênica. A cepa de Pichia pastoris Glycoswitch tem a maquinaria para expressar e secretar altas quantidades de enzima com glicosilação humanizada. Nesse trabalho, descrevemos o processo genético para expressar a ScASNaseII no meio extracelular pela P. pastoris Glycoswitch, e também os parâmetros bioquímicos, perfil cinético, citotoxicidade contra células leucêmicas e a interferência da glicosilação na atividade da enzima obtida. Nossos dados mostram que a cepa aplicada foi capaz de expressar ScASNaseII no meio extracelular passível de purificação de proteínas contaminantes com apenas um passo cromatográfico. A atividade específica para asparagina foi 218,2 UI/mg e a atividade glutaminásica representou 3,1% da atividade asparaginásica. Os parâmetros cinéticos foram KM = 120,5 µM e a eficiência catalítica de 3,8 x 105 M-1s-1. Análises por meio de gel nativo sugerem uma conformação tetramérica de aproximadamente 150 kDa. Essa é uma nova estratégia de produzir essa enzima de forma extracelular, com mais facilidade de purificação e com melhores propriedades biotecnológicas


L-asparaginase is an efficient inhibitor of tumor development, used in chemotherapy sessions against acute lymphoblastic leukemia (ALL) tumor cell; its use results in 90% complete remission of the disease in treated patients. Saccharomyces cerevisiae's L-asparaginase II (ScASNaseII) has a high potential to overcome the side effects of bacteria L-asparaginase, but the endogenous production of it results in hypermannosylated immunogenic enzyme. However, Pichia pastoris Glycoswitch strain has the machinery to express and secrete high quantity of the enzyme and with humanized glycosylation. Here we describe the genetic process to acquire the ScASNaseII in the extracellular medium expressed by P. pastoris Glycoswitch, and the biochemical properties of the resultant enzyme, kinetic profile, cytotoxicity against ALL cell line and the interference of glycosylation in its activity. Our data show that the strain employed is able to express extracellular asparaginase active and possible to be purified of contaminant proteins using a single chromatographic step. The specific activity using asparagine was 218.2 IU.mg-1 and the glutaminase activity represents 3.1% of its asparaginase activity. The kinetics parameters were KM=120.5 µM and a catalytic efficiency of 3.8x105 M-1s-1. The Native-PAGE suggested a tetrameric protein conformation, with approximately 150 kDa. This is a novel strategy to produce this enzyme extracellularly, easier to purify and with better biotechnological properties


Subject(s)
Pichia/isolation & purification , Asparaginase/analysis , Saccharomyces cerevisiae/isolation & purification , Glycosylation , Recombinant Proteins , Precursor Cell Lymphoblastic Leukemia-Lymphoma/diagnosis
13.
Araçatuba; s.n; 2019. 115 p. graf, ilus, tab.
Thesis in Portuguese | LILACS, BBO | ID: biblio-1051141

ABSTRACT

A redução da reatividade vascular à fenilefrina (PE) em aorta de ratas espontaneamente hipertensas (SHR) ao final da prenhez é dependente de maior produção e/ou maior biodisponibilidade de óxido nítrico (NO), consequente do aumento da fosforilação da enzima óxido nítrico sintase endotelial (eNOS) via PI3K/Akt. A glicosilação do tipo N-acetil-glucosamina (O-GlcNAc) é uma modificação pós-traducional que compete com a fosforilação pelos mesmos sítios de ligação nas proteínas. A O-GlcNAcilação da eNOS em serina1177 leva a redução da sua atividade enquanto a fosforilação leva a sua ativação. Além destes mecanismos, a interação da eNOS com outras proteínas é capaz de regular positiva ou negativamente a sua atividade. O objetivo deste trabalho foi analisar possíveis alterações nos mecanismos de modificação pós-traducional que controlam a ativação da eNOS os quais poderiam contribuir para maior ativação e maior biodisponibilidade de NO observada em artérias de ratas prenhes. Foram avaliados o conteúdo proteico O-GlcNAc e também expressão das enzimas que participam desta modificação, O-GlcNAc transferase (OGT) e O-GlcNAcase (OGA) por Western Blotting e a atividade da OGA por ensaio bioquímico em aorta e em artéria mesentérica (2º ou 3º ramo) de ratas não prenhes (NP) e prenhes (P), normotensas (Wistar) e SHR. Ensaios de Western Blotting foram realizados também para análise da expressão das seguintes proteínas: Cav-1, p-Cav-1, CaM e Hsp90. Realizamos a contagem do número de cavéolas endoteliais da aorta e da artéria mesentérica na presença ou ausência da metil-ß-ciclodextrina (dextrina, 10 mmol/L) por microscopia eletrônica. Em estudos funcionais, avaliamos a participação da enzima OGA, pela inibição com PugNAc (100 µmol/L) e das cavéolas, utilizando um desorganizador de cavéolas, a dextrina (10 ou 20 mmol/L), na menor reatividade vascular à PE observada em aortas de ratas P. Observamos que o conteúdo de proteínas O-GlcNAciladas estava diminuído em aorta e em leito mesentérico de ratas Wistar P e SHR P. Apesar da expressão da OGT e da OGA não estar alterada, a atividade da OGA foi aumentada em aorta e leito mesentérico de ratas Wistar P, mas, encontra-se diminuída em aorta e aumentada em leito mesentérico de SHP P. A incubação com PugNAc reverteu a reduzida reatividade à PE em aorta e artéria mesentérica de ratas Wistar P mas este efeito não foi observado em vasos SHR P, demonstrando que a OGA parece ter um papel importante na redução da O-GlcNAcilação de proteínas vasculares em Wistar P. Em vasos incubados com PugNAc, a remoção do endotélio ou a incubação com L-NAME, não alterou significativamente a reatividade à PE. Juntos estes resultados sugerem que a maior atividade da eNOS observada em vasos de Wistar P, fica prejudicada na presença do PugNAc, e depende da atividade da OGA. Como não houve alteração da resposta contrátil à PE em vasos de SHR P incubados com PugNAc, possivelmente um mecanismo diferente, envolvendo a menor atividade da OGT, ocorre nestas artérias para a redução da O-GlcNAcilação da eNOS. A desorganização das cavéolas por meio da dextrina causou aumento de contração à PE e redução de potência da ACh em aortas de Wistar NP e SHR NP, porém não houve alteração em aortas de ratas Wistar P e SHR P. A dextrina não alterou o número de cavéolas em artérias de Wistar P e SHR P quando comparado com ratas NP. SHR NP apresentam um reduzido número de cavéolas das aortas em relação a Wistar NP bem como expressão reduzida de Cav-1, p-Cav-1 e CaM. A prenhez não foi capaz de alterar a expressão da Cav-1, CaM e Hsp90 em aorta e leito mesentérico de ratas normotensas e hipertensas. Estes resultados sugerem que a prenhez não altera a expressão das proteínas Cav-1, CaM e Hsp90 e possivelmente a interação com a eNOS em aorta e artérias mesentéricas de ratas normotensas e hipertensas. Em conclusão, entre os mecanismos estudados de modificação pós-traducional da eNOS, a redução da O-GlcNAcilação da eNOS, por mecanismos que envolvem a atividade da OGA e possivelmente da OGT, favoreceria a fosforilação da eNOS e consequente maior biodisponibilidade de NO, contribuindo desta forma para modulação da resposta contrátil da PE nas artérias de ratas P(AU)


Reduction of vascular reactivity to phenylephrine (PE) in aorta of spontaneously hypertensive rats (SHR) at the end of pregnancy is dependent on higher production and/or higer bioavailability of nitric oxide (NO), as a consequence of increased endothelial nitric oxide synthase enzyme (eNOS) phosphorylation, by PI3K/Akt. Glycosylation with O-linked N-acetylglucosamine (O-GlcNAc) is a post-translational modification that competes with phosphorylation by the same binding sites in proteins. O-GlcNAcylation of eNOS on serine site leads to a reduction in its activity while eNOS phosphorylation leads to its activation. In addition to these mechanisms, the interaction of eNOS with other proteins is able to regulate positively or negatively its activity. The objective of this study was to analyze possible changes in the mechanisms of post-translational modification that control the eNOS activation, which could contribute to its the greater activation and greater bioavailability of NO observed in arteries of pregnant rats. The O-GlcNAc-protein content and also the enzymes expression that participate in this modification, O-GlcNAc transferase (OGT) and O-GlcNAcase (OGA) was assessed by Western Blotting, and OGA activity were evaluated by biochemical assay in the aorta and in the artery mesenteric (2nd or 3rd branch) of non-pregnant (NP) and pregnant (P), normotensive rats (Wistar) and SHR. Western Blotting assays were also performed for expression analysis of the following proteins: Cav-1, p-Cav-1, CaM and Hsp90. We performed the counting of the number of endothelial caveolae in the aorta and the mesenteric artery in the presence or absence of methyl-ß-cyclodextrin (dextrin, 10 mmol/L) by electronic microscopy. In functional studies, we evaluated the participation of the OGA enzyme, by inhibition with PugNAc (100 µmol/L) and of the caveolae, using a caveolae disassembler, dextrin (10 or 20 mmol/L), in the reduced vascular reactivity observed in aortas or mesenteric arteries of P rats. We observed that the content of O-GlcNAcylated proteins was decreased in the aorta and in the mesenteric bed of Wistar P and SHR P rats. Although OGT and OGA expression is not altered, OGA activity was increased in the aorta and mesenteric bed of Wistar P rats but was decreased in the aorta and increased in the mesenteric bed of SHP P. Incubation with PugNAc reversed the reduced reactivity to PE in the aorta and mesenteric artery of Wistar P but this effect was not observed in SHR P arteries, demonstrating that OGA appears to play an important role in reducing O-GlcNAcylation of vascular proteins in Wistar P. In arteries incubated with PugNAc, endothelial removal or incubation with L-NAME did not significantly alter reactivity to PE. Together, these results suggest that the greater eNOS activity observed in Wistar P vessels was impaired in the presence of PugNAc, and it depends on OGA activity. As there was no change in the contractile response to PE in SHR P arteries incubated with PugNAc, possibly a different mechanism, involving the lower activity of OGT, occurs in these vessels for the reduction of O-GlcNAcylation of eNOS. Dextrin caused increased contraction of PE and decreased ACh potency in Wistar NP and SHR NP aortas, but there was no change in aortas of Wistar P and SHR P. Dextrin did not alter the number of cavelae in Wistar P and SHR P arteries compared to NP rats. SHR NP showed a lower number of caveolae than to NP Wistar as well reduced expression of Cav-1 and CaM. Pregnancy was not able to alter the expression of Cav-1, CaM and Hsp90 in the aorta and mesenteric bed of normotensive and hypertensive rats. These results suggest that pregnancy does not alter the expression of Cav-1, CaM and Hsp90 proteins and possibly interaction with eNOS in the aorta and mesenteric arteries of normotensive and hypertensive rats. In conclusion, among the studied mechanisms of post-translational modification of eNOS, the reduction of O-GlcNAcylation of eNOS, by mechanisms that involve OGA activity and possibly OGT, would favor eNOS phosphorylation and consequent greater NO bioavailability, contributing in this way for modulation of the contractile response to PE in the arteries of P rats(AU)


Subject(s)
Animals , Female , Pregnancy , Aorta , Nitric Oxide Synthase , Hypertension , Glycosylation , Calmodulin , Rats, Wistar , HSP90 Heat-Shock Proteins , Caveolin 1 , Mesenteric Arteries
14.
Rio de Janeiro; s.n; 2019. xiv, 152 p. ilus.
Thesis in Portuguese | LILACS | ID: biblio-1049943

ABSTRACT

Celulases fúngicas têm sido usadas para degradar a biomassa lignocelulósica para a produção de bioetanol. Celulases industriais como Cel7A de Trichoderma reesei (TrCel7A) são críticas neste processo. A compreensão da estrutura e dinâmica é crucial para a reengenharia da atividade celulolítica. Esta enzima é formada por dois domínios ligados por um linker flexível e altamente glicosilado. No entanto, a flexibilidade do linker tem dificultado a determinação da estrutura completa da Cel7A. Assim, na ausência de dados experimentais de alta resolução, aplicamos a modelagem integrativa para construir um modelo da enzima completa. Em seguida, estudamos os efeitos da glicosilação na estrutura e dinâmica da apo TrCel7A por meio de simulações. A análise da dinâmica essencial mostrou que a O-glicosilação no linker levou à estabilização da dinâmica global da proteína. Os glicanos O-ligados parecem restringir a distribuição dos ângulos diedros desta região, selecionando conformações mais alongadas. Além da flexibilidade reduzida, os movimentos interdomínios funcionais foram preservados no sistema glicosilado. Em contraste, observamos grande plasticidade conformacional na ausência de glicosilação, mas os domínios funcionais frequentemente colapsaram. Nós relatamos aqui evidências de que a flexibilidade dirigida no linker de Cel7A por mutações pontuais, incluindo modificações de sítios de glicosilação, poderia ser uma estratégia promissora para melhorar a atividade da celulase. (AU)


Subject(s)
Humans , Trichoderma , Glycosylation , Mutagenesis, Insertional , Cellulases
15.
Rev. Soc. Argent. Diabetes ; 53(1): 32-33, Ene.-Abr. 2019.
Article in Spanish | LILACS | ID: biblio-1021894

ABSTRACT

La interacción entre uroepitelio y uropatógeno, base de la patogenia de las infecciones del tracto urinario (ITUs), puede derivar en la eliminación bacteriana por parte de la célula huésped o la invasión y multiplicación bacteriana. Dentro de la célula huésped los uropatógenos pueden perturbar las defensas y resistir el tratamiento antibiótico. En pacientes con diabetes, especialmente con enfermedad renal por diabetes, se ha demostrado una reducción de la capacidad de inhibición de la adherencia bacteriana al uroepitelio, por ende mayor posibilidad de invasión bacteriana. La glicosilación de todos los elementos del sistema inmune, incluida la menor liberación de factores como las interleuquinas a nivel urinario y la alteración del vaciamiento vesical por neuropatía autonómica, favorecen el desarrollo de este tipo de infecciones


Interaction between urothelium and uropathogen, the basis of the pathogenesis of urinary tract infections (UTIs), can lead to bacterial elimination by the host cell or bacterial invasion and multiplication. Inside the host cell, uropathogens can impair defenses and resist antibiotic treatment. In patients with diabetes, especially diabetes-related kidney disease, a reduction in the inhibition capacity of bacterial adherence to the urothelium has been demonstrated; therefore, a highest chance of bacterial invasion. The glycosylation of all elements of the immune system, including the lower release of factors such as interleukins at the urinary level and the impairment of bladder emptying by autonomic neuropathy, enhance the development of this type of infections


Subject(s)
Glycosylation , Diabetes Mellitus , Uropathogenic Escherichia coli , Uroplakins
16.
Actual. osteol ; 14(3): 205-218, sept. - dic. 2018. ilus., graf.
Article in Spanish | LILACS | ID: biblio-1052695

ABSTRACT

La diabetes es una enfermedad crónica asociada con importantes comorbilidades. El sistema esquelético parece ser un objetivo adicional de daño mediado por diabetes. Se acepta que la diabetes tipo 1 y tipo 2 se asocian con un mayor riesgo de fractura ósea. Varios estudios han demostrado que los cambios metabólicos causados por la diabetes pueden influir en el metabolismo óseo disminuyendo la calidad y la resistencia del hueso. Sin embargo, los mecanismos subyacentes no se conocen por completo pero son multifactoriales y, probablemente, incluyen los efectos de la obesidad, hiperglucemia, estrés oxidativo y acumulación de productos finales de glicosilación avanzada. Estos darían lugar a un desequilibrio de varios procesos y sistemas: formación de hueso, resorción ósea, formación y entrecruzamiento de colágeno. Otros factores adicionales como la hipoglucemia inducida por el tratamiento, ciertos medicamentos antidiabéticos con un efecto directo sobre el metabolismo óseo y mineral, así como una mayor propensión a las caídas, contribuirían al aumento del riesgo de fracturas en pacientes con diabetes mellitus. Esta revisión tiene como objetivo describir los mecanismos fisiopatológicas subyacentes a la fragilidad ósea en pacientes diabéticos. (AU)


Diabetes is a chronic disease associated with important comorbidities. The skeletal system seems to be an additional target of diabetes mediated damage. It is accepted that type 1 and type 2 diabetes are associated with an increased risk of bone fracture. Several studies have shown that metabolic changes caused by diabetes can influence bone metabolism by decreasing bone quality and resistance. However, the underlying mechanisms are not completely known but they are multifactorial and probably include the effects of obesity, hyperglycemia, oxidative stress and accumulation of advanced glycosylation end products. These would lead to an imbalance of several processes and systems: bone formation, bone resorption, formation and collagen crosslinking. Other additional factors such as treatment-induced hypoglycemia, certain antidiabetic medications with a direct effect on bone and mineral metabolism, as well as an increased propensity for falls, would contribute to the increased risk of fractures in patients with diabetes mellitus. This review aims to describe the pathophysiological mechanisms underlying bone fragility in diabetic patients. (AU)


Subject(s)
Humans , Male , Female , Middle Aged , Osteogenesis Imperfecta/physiopathology , Diabetes Mellitus/physiopathology , Osteogenesis Imperfecta/metabolism , Osteogenesis Imperfecta/drug therapy , Osteoporosis/diagnosis , Bone and Bones/metabolism , Glycosylation , Risk Factors , Oxidative Stress , Diabetes Mellitus/metabolism , Diabetes Mellitus/epidemiology , Diabetes Mellitus, Type 1/complications , Diabetes Mellitus, Type 2/complications , Fractures, Bone/complications , Fractures, Bone/prevention & control , Hyperglycemia/complications , Hypoglycemia/chemically induced , Hypoglycemic Agents/adverse effects , Obesity/complications
17.
Medicina (Bogotá) ; 40(1(120)): 113-113, Ene-Mar, 2018.
Article in Spanish | LILACS | ID: biblio-910095

ABSTRACT

Introducción: La glicosilación es una modificación postraduccional que tiene un papel protagónico en los procesos de comunicación célula-célula, célula-matrix extracelular, por lo tanto modula la interacción con antígenos y el sistema inmune. La inmunoglobulina G (IgG) tiene un sitio de N-glicosilación en la posición 297. Los sacáridos terminales de este glicano impactan la función efectora de la proteína, dirigiendo la respuesta hacia un efecto proinflamatorio o antiinflamatorio. Maverakis. et al (2015), propusieron que cada enfermedad autoinmune tiene una única marca de glicanos, especialmente en las diferentes clases de inmunoglobulinas (teoría de la alteración de los glicanos en la autoinmunidad).


Subject(s)
Antiphospholipid Syndrome , Autoimmunity , Glycosylation , Immunoglobulin G
18.
Article in English | WPRIM | ID: wpr-690636

ABSTRACT

Nonalcoholic fatty liver disease (NAFLD) is a major public health issue worldwide. Immunoglobulin G (IgG) N-glycans are associated with risk factors for NAFLD, such as obesity and diabetes. A cross-sectional study involving 500 Han Chinese adults recruited from a community in Beijing was carried out to explore the association between IgG N-glycans and NAFLD. IgG N-glycosylation was significantly associated with NAFLD, with the disease showing a negative correlation with galactosylation (GP14, GP14n, and G2n), positive correlation with fucosylation (FBG2n/G2n), and positive correlation with bisecting N-acetylglucosamine (GlcNAc) [FBG2n/FG2n and FBG2n/(FG2n+FBG2n)], after controlling age, gender, and prevalence of obesity, type 2 diabetes mellitus, hypertension, and hyperlipidemia. In other words, the present study showed a possible association between NAFLD and the loss of galactose and elevations of fucose and bisecting GlcNAc. Aberrant IgG glycosylation might therefore be a potential biomarker for the primary or secondary prevention of NAFLD.


Subject(s)
Biomarkers , Blood , China , Cross-Sectional Studies , Diabetes Mellitus, Type 2 , Blood , Female , Glycosylation , Humans , Immunoglobulin G , Blood , Male , Middle Aged , Non-alcoholic Fatty Liver Disease , Blood , Obesity , Blood , Odds Ratio , Polysaccharides , Blood , Risk Factors
20.
Article in English | WPRIM | ID: wpr-715661

ABSTRACT

BACKGROUND: Liver biopsies have been partially replaced by noninvasive methods for assessing liver fibrosis. We explored the usefulness of four novel biomarkers, enhanced liver fibrosis (ELF), glycosylation isomer of Mac-2 binding protein (M2BPGi), galectin-3, and soluble suppression of tumorigenicity 2 (sST2), in association with liver fibrosis. METHODS: ELF, M2BPGi, galectin-3, and sST2 were assayed in 173 patients with chronic liver diseases. The results were analyzed according to fibrosis grade (F0/1, F2, and F3/4) by transient elastography (TE). RESULTS: ELF, M2BPGi, galectin-3, and sST2 values differed significantly according to TE grade; ELF and M2BPGi values were higher in F2 and F3/4 than in F0/1 (P≤0.001, all), sST2 values were higher in F3/4 than in F0/1 and F2 (P < 0.05), and galectin-3 values were higher in F3/4 than in F0/1 (P=0.0036). ELF and M2BPGi showed good TE fibrosis detection performance (area under the curves [AUC], 0.841 and 0.833 for ≥F2; and 0.837 and 0.808 for ≥F3). The sensitivity and specificity for predicting TE grade F≥2 were 84.1% and 76.7% for ELF and 63.6% and 91.5% for M2BPGi. CONCLUSIONS: This is the first study to compare the liver fibrosis assessment of four novel biomarkers: ELF, M2BPGi, galectin-3, and sST2. The biomarkers varied significantly according to TE grade, and each biomarker showed a different trend. ELF and M2BPGi seem to have comparable good performance for detecting liver fibrosis.


Subject(s)
Biomarkers , Biopsy , Carrier Proteins , Elasticity Imaging Techniques , Fibrosis , Galectin 3 , Glycosylation , Humans , Liver Cirrhosis , Liver Diseases , Liver , Sensitivity and Specificity
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