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1.
J Med Virol ; 93(5): 3113-3121, 2021 May.
Artigo em Inglês | MEDLINE | ID: mdl-33570194

RESUMO

The clinical symptoms of community-acquired pneumonia (CAP) and coronavirus disease 2019 (COVID-19)-associated pneumonia are similar. Effective predictive markers are needed to differentiate COVID-19 pneumonia from CAP in the current pandemic conditions. Copeptin, a 39-aminoacid glycopeptide, is a C-terminal part of the precursor pre-provasopressin (pre-proAVP). The activation of the AVP system stimulates copeptin secretion in equimolar amounts with AVP. This study aims to determine serum copeptin levels in patients with CAP and COVID-19 pneumonia and to analyze the power of copeptin in predicting COVID-19 pneumonia. The study consists of 98 patients with COVID-19 and 44 patients with CAP. The basic demographic and clinical data of all patients were recorded, and blood samples were collected. The receiver operating characteristic (ROC) curve was generated and the area under the ROC curve (AUC) was measured to evaluate the discriminative ability. Serum copeptin levels were significantly higher in COVID-19 patients compared to CAP patients (10.2 ± 4.4 ng/ml and 7.1 ± 3.1 ng/ml; p < .001). Serum copeptin levels were positively correlated with leukocyte, neutrophil, and platelet count (r = -.21, p = .012; r = -.21, p = .013; r = -.20, p = .018; respectively). The multivariable logistic regression analysis revealed that increased copeptin (odds ratio [OR] = 1.183, 95% confidence interval [CI], 1.033-1.354; p = .015) and CK-MB (OR = 1.052, 95% CI, 1.013-1.092; p = .008) levels and decreased leukocyte count (OR = 0.829, 95% CI, 0.730-0.940; p = .004) were independent predictors of COVID-19 pneumonia. A cut-off value of 6.83 ng/ml for copeptin predicted COVID-19 with a sensitivity of 78% and a specificity of 73% (AUC: 0.764% 95 Cl: 0.671-0.856, p < .001). Copeptin could be a promising and useful biomarker to be used to distinguish COVID-19 patients from CAP patients.


Assuntos
/diagnóstico , Glicopeptídeos/sangue , Pneumonia Bacteriana/diagnóstico , Adulto , Idoso , Idoso de 80 Anos ou mais , Biomarcadores/sangue , Infecções Comunitárias Adquiridas , Feminino , Glicopeptídeos/metabolismo , Humanos , Modelos Logísticos , Masculino , Pessoa de Meia-Idade
2.
Eur J Endocrinol ; 183(6): 669-676, 2020 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-33112256

RESUMO

Context: Obesity and cardiometabolic diseases are associated with higher long-term glucocorticoid levels, measured as scalp hair cortisol (HairF) and cortisone (HairE). Cardiometabolic diseases have also been associated with copeptin, a stable surrogate marker for the arginine-vasopressin (AVP) system. Since AVP is, together with corticotropin-releasing hormone (CRH) an important regulator of the hypothalamic-pituitary adrenal axis (HPA axis), we hypothesize that AVP contributes to chronic hypercortisolism in obesity. Objective: To investigate whether copeptin levels are associated with Higher HairF and HairE levels in obesity. Design: A cross-sectional study in 51 adults with obesity (BMI ≥30 kg/m2). Methods: Associations and interactions between copeptin, HairF, HairE, and cardiometabolic parameters were cross-sectionally analyzed. Results: Copeptin was strongly associated with BMI and waist circumference (WC) (rho = 0.364 and 0.530, P = 0.008 and <0.001, respectively), also after correction for confounders. There were no associations between copeptin and HairF or HairE on a continuous or dichotomized scale, despite correction for confounders. Conclusion: In patients with obesity, AVP seems not a major contributor to the frequently observed high cortisol levels. Other factors which stimulate the HPA axis or affect cortisol synthesis or breakdown may be more important than the influence of AVP on long-term glucocorticoid levels in obesity.


Assuntos
Cortisona/metabolismo , Síndrome de Cushing/etiologia , Glicopeptídeos/metabolismo , Hidrocortisona/metabolismo , Obesidade/metabolismo , Adulto , Arginina Vasopressina/metabolismo , Biomarcadores/metabolismo , Índice de Massa Corporal , Hormônio Liberador da Corticotropina/metabolismo , Estudos Transversais , Feminino , Glucocorticoides/metabolismo , Cabelo/química , Humanos , Sistema Hipotálamo-Hipofisário/metabolismo , Masculino , Obesidade/complicações , Sistema Hipófise-Suprarrenal/metabolismo
3.
PLoS One ; 15(8): e0238288, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32857795

RESUMO

BACKGROUND: The aim of this study was to evaluate the value of copeptin in predicting mortality including both short-term and long-term mortality in patients with acute coronary syndrome (ACS). METHODS: Potential studies were searched and selected through PubMed, Embase and Cochrane databases up to December 2019. The predictive performance was evaluated by the pooled sensitivity and specificity, and summary receiver operating characteristic curves. Cochran's Q test and I2 index were used to assess between-study heterogeneity, and Deek's test and funnel plots were used to assess publication bias. RESULTS: Total six studies comprising 2269 patients were included in this meta-analysis. The area under the receiver operating characteristic curve of copeptin in predicting mortality in patients with ACS was 0.73 (95% CI: 0.69-0.77). The pooled sensitivity and specificity of copeptin were 0.77 (95% CI: 0.59-0.89) and 0.60 (95% CI: 0.47-0.71), respectively. Significant between-study heterogeneity was identified in both sensitivity (P = 0.01; I2 = 69.76%) and specificity (P<0.001; I2 = 97.32%) among the six included studies. The meta-regression analysis indicated that the number of study centers was significantly associated with the heterogeneity of sensitivity (P = 0.03), whereas the study design (P = 0.03) and duration of follow-up (P<0.001) were significantly associated with the heterogeneity of specificity. CONCLUSIONS: Copeptin has acceptable prognostic value for mortality in patients with ACS. Further studies based on multimarker strategy are needed to evaluate the prognostic value of copeptin for ACS in conjunction with other well-established biomarkers.


Assuntos
Síndrome Coronariana Aguda/metabolismo , Glicopeptídeos/metabolismo , Síndrome Coronariana Aguda/mortalidade , Humanos , Prognóstico
4.
Nature ; 584(7820): 291-297, 2020 08.
Artigo em Inglês | MEDLINE | ID: mdl-32728216

RESUMO

The majority of therapies that target individual proteins rely on specific activity-modulating interactions with the target protein-for example, enzyme inhibition or ligand blocking. However, several major classes of therapeutically relevant proteins have unknown or inaccessible activity profiles and so cannot be targeted by such strategies. Protein-degradation platforms such as proteolysis-targeting chimaeras (PROTACs)1,2 and others (for example, dTAGs3, Trim-Away4, chaperone-mediated autophagy targeting5 and SNIPERs6) have been developed for proteins that are typically difficult to target; however, these methods involve the manipulation of intracellular protein degradation machinery and are therefore fundamentally limited to proteins that contain cytosolic domains to which ligands can bind and recruit the requisite cellular components. Extracellular and membrane-associated proteins-the products of 40% of all protein-encoding genes7-are key agents in cancer, ageing-related diseases and autoimmune disorders8, and so a general strategy to selectively degrade these proteins has the potential to improve human health. Here we establish the targeted degradation of extracellular and membrane-associated proteins using conjugates that bind both a cell-surface lysosome-shuttling receptor and the extracellular domain of a target protein. These initial lysosome-targeting chimaeras, which we term LYTACs, consist of a small molecule or antibody fused to chemically synthesized glycopeptide ligands that are agonists of the cation-independent mannose-6-phosphate receptor (CI-M6PR). We use LYTACs to develop a CRISPR interference screen that reveals the biochemical pathway for CI-M6PR-mediated cargo internalization in cell lines, and uncover the exocyst complex as a previously unidentified-but essential-component of this pathway. We demonstrate the scope of this platform through the degradation of therapeutically relevant proteins, including apolipoprotein E4, epidermal growth factor receptor, CD71 and programmed death-ligand 1. Our results establish a modular strategy for directing secreted and membrane proteins for lysosomal degradation, with broad implications for biochemical research and for therapeutics.


Assuntos
Espaço Extracelular/metabolismo , Lisossomos/metabolismo , Proteínas de Membrana/metabolismo , Proteólise , Proteínas Recombinantes de Fusão/metabolismo , Animais , Anticorpos/química , Anticorpos/metabolismo , Antígenos CD/metabolismo , Apolipoproteína E4/metabolismo , Antígeno B7-H1/metabolismo , Sistemas CRISPR-Cas , Linhagem Celular , Receptores ErbB/metabolismo , Feminino , Glicopeptídeos/síntese química , Glicopeptídeos/metabolismo , Humanos , Ligantes , Proteínas de Membrana/química , Camundongos , Domínios Proteicos , Transporte Proteico , Receptor IGF Tipo 2/metabolismo , Receptores da Transferrina/metabolismo , Proteínas Recombinantes de Fusão/síntese química , Proteínas Recombinantes de Fusão/química , Solubilidade , Especificidade por Substrato
5.
Sci Rep ; 10(1): 2708, 2020 02 17.
Artigo em Inglês | MEDLINE | ID: mdl-32066783

RESUMO

Prostate-specific antigen (PSA) is the most frequently used biomarker for the screening of prostate cancer. Understanding the structure of cancer-specific glycans can help us improve PSA assay. In the present study, we analysed the glycans of PSA obtained from culture medium containing cancer tissue-originated spheroids (CTOS) which have similar characteristics as that of the parent tumour to explore the new candidates for cancer-related glycoforms of PSA. The glycan profile of PSA from CTOS was determined by comparing with PSA from normal seminal plasma and cancer cell lines (LNCaP and 22Rv1) using lectin chromatography and mass spectrometry. PSA from CTOS was mostly sialylated and the content of Wisteria floribunda agglutinin reactive glycan (LacdiNAc) was similar to that of PSA derived from seminal plasma and 22Rv1. Conversely, concanavalin A (Con A)-unbound PSA was definitely detected from the three cancer origins but was almost negligible in seminal PSA. Two novel types of PSA were elucidated in the Con A-unbound fraction: one is a high molecular weight PSA with highly branched N-glycans, and the other is a low molecular weight PSA without N-glycans. Furthermore, the existence of Lewis X antigen group on PSA was indicated. These PSAs will be candidates for new cancer-related markers.


Assuntos
Biomarcadores Tumorais/metabolismo , Polissacarídeos/química , Antígeno Prostático Específico/metabolismo , Neoplasias da Próstata/diagnóstico , Processamento de Proteína Pós-Traducional , Esferoides Celulares/metabolismo , Biomarcadores Tumorais/química , Sequência de Carboidratos , Linhagem Celular Tumoral , Cromatografia de Afinidade , Concanavalina A/química , Meios de Cultivo Condicionados/química , Glicopeptídeos/química , Glicopeptídeos/metabolismo , Glicosilação , Humanos , Antígenos CD15/química , Antígenos CD15/metabolismo , Masculino , Lectinas de Plantas/química , Polissacarídeos/isolamento & purificação , Polissacarídeos/metabolismo , Próstata/metabolismo , Próstata/patologia , Antígeno Prostático Específico/química , Neoplasias da Próstata/metabolismo , Neoplasias da Próstata/patologia , Receptores de N-Acetilglucosamina/química , Sêmen/química , Espectrometria de Massas por Ionização e Dessorção a Laser Assistida por Matriz , Esferoides Celulares/química , Esferoides Celulares/patologia
6.
ACS Appl Mater Interfaces ; 12(8): 9814-9823, 2020 Feb 26.
Artigo em Inglês | MEDLINE | ID: mdl-32011110

RESUMO

In this work, a magnetic covalent organic framework (COF) with inherent hydrophilicity (denoted mCTpBD) was synthesized through interface deposition of a hydrophilic COF shell on amino group-functionalized magnetite particles via the reaction between a carboxyl group-containing monomer and benzidine. Thanks to the superior hydrophilicity, appropriate porous structure, and easy magnetic separation, the resulting mCTpBD exhibited excellent performance in conveniently enriching glycopeptides from standard samples with a high sensitivity of 0.5 fmol µL-1 and strong size-exclusion effect of up to 1:1000 (w/w). Furthermore, by using the mCTpBD adsorbent, endogenous glycopeptides in saliva of healthy people and patients with inflammatory bowel disease were successfully enriched and identified by the combined liquid chromatography-mass spectrometry/mass spectrometry technology, which indicates a promising prospective of core-shell magnetic composite microspheres with a hydrophilic COF shell in glycoproteomics research.


Assuntos
Glicopeptídeos/metabolismo , Doenças Inflamatórias Intestinais/metabolismo , Nanopartículas de Magnetita/química , Estruturas Metalorgânicas/química , Saliva/metabolismo , Proteínas e Peptídeos Salivares/metabolismo , Humanos
7.
Sci Rep ; 10(1): 318, 2020 01 15.
Artigo em Inglês | MEDLINE | ID: mdl-31941975

RESUMO

Protein glycosylation is known to be involved in biological progresses such as cell recognition, growth, differentiation, and apoptosis. Fucosylation of glycoproteins plays an important role for structural stability and function of N-linked glycoproteins. Although many of biological and clinical studies of protein fucosylation by fucosyltransferases has been reported, structural classification of fucosylated N-glycoproteins such as core or outer isoforms remains a challenge. Here, we report for the first time the classification of N-glycopeptides as core- and outer-fucosylated types using tandem mass spectrometry (MS/MS) and machine learning algorithms such as the deep neural network (DNN) and support vector machine (SVM). Training and test sets of more than 800 MS/MS spectra of N-glycopeptides from the immunoglobulin gamma and alpha 1-acid-glycoprotein standards were selected for classification of the fucosylation types using supervised learning models. The best-performing model had an accuracy of more than 99% against manual characterization and area under the curve values greater than 0.99, which were calculated by probability scores from target and decoy datasets. Finally, this model was applied to classify fucosylated N-glycoproteins from human plasma. A total of 82N-glycopeptides, with 54 core-, 24 outer-, and 4 dual-fucosylation types derived from 54 glycoproteins, were commonly classified as the same type in both the DNN and SVM. Specifically, outer fucosylation was dominant in tri- and tetra-antennary N-glycopeptides, while core fucosylation was dominant in the mono-, bi-antennary and hybrid types of N-glycoproteins in human plasma. Thus, the machine learning methods can be combined with MS/MS to distinguish between different isoforms of fucosylated N-glycopeptides.


Assuntos
Fucose/análise , Cadeias gama de Imunoglobulina/metabolismo , Aprendizado de Máquina , Espectrometria de Massas em Tandem/métodos , Sequência de Aminoácidos , Sequência de Carboidratos , Cromatografia Líquida de Alta Pressão , Glicopeptídeos/análise , Glicopeptídeos/química , Glicopeptídeos/metabolismo , Glicosilação , Humanos
8.
Mol Cell Proteomics ; 19(3): 529-539, 2020 03.
Artigo em Inglês | MEDLINE | ID: mdl-31924694

RESUMO

Glycosylation is a common modification of proteins and critical for a wide range of biological processes. Differences in protein glycosylation between sexes have already been observed in humans, nematodes and trematodes, and have recently also been reported in the rice pest insect Nilaparvata lugens Although protein N-glycosylation in insects is nowadays of high interest because of its potential for exploitation in pest control strategies, the functionality of differential N-glycosylation between sexes is yet unknown. In this study, therefore, the occurrence and role of sex-related protein N-glycosylation in insects were examined. A comprehensive investigation of the N-glycosylation sites from the adult stages of N. lugens was conducted, allowing a qualitative and quantitative comparison between sexes at the glycopeptide level. N-glycopeptide enrichment via lectin capturing using the high mannose/paucimannose-binding lectin Concanavalin A, or the Rhizoctonia solani agglutinin which interacts with complex N-glycans, resulted in the identification of over 1300 N-glycosylation sites derived from over 600 glycoproteins. Comparison of these N-glycopeptides revealed striking differences in protein N-glycosylation between sexes. Male- and female-specific N-glycosylation sites were identified, and some of these sex-specific N-glycosylation sites were shown to be derived from proteins with a putative role in insect reproduction. In addition, differential glycan composition between males and females was observed for proteins shared across sexes. Both lectin blotting experiments as well as transcript expression analyses with complete insects and insect tissues confirmed the observed differences in N-glycosylation of proteins between sexes. In conclusion, this study provides further evidence for protein N-glycosylation to be sex-related in insects. Furthermore, original data on N-glycosylation sites of N. lugens adults are presented, providing novel insights into planthopper's biology and information for future biological pest control strategies.


Assuntos
Glicopeptídeos/metabolismo , Hemípteros/metabolismo , Proteínas de Insetos/metabolismo , Caracteres Sexuais , Animais , Feminino , Trato Gastrointestinal/metabolismo , Glicosilação , Cabeça , Masculino , Ovário/metabolismo , Reprodução , Testículo/metabolismo
9.
Am J Physiol Renal Physiol ; 318(3): F702-F709, 2020 03 01.
Artigo em Inglês | MEDLINE | ID: mdl-31961713

RESUMO

Alcohol consumption influences sodium-water homeostasis. However, the effect of alcohol on vasopressin levels is controversial. The aim of the present study was to evaluate physiological changes of alcohol consumption on the stable vasopressin surrogate marker copeptin. In addition, we aimed at investigating the effect of additional sodium and/or water consumption on plasma sodium, osmolality, and copeptin levels. Ten healthy men underwent four interventions in random order: 1) beer consumption only, 2) beer consumption with additional water, 3) beer consumption with additional stock, or 4) water consumption only. Fluid consumption was equal between interventions and calculated to reach a blood alcohol concentration of 0.8‰ in the beer interventions. Blood and urinary samples were taken at six time points over the observation period of 720 min. The primary end point was the mean difference in copeptin levels 90 min after the start of fluid consumption, which showed no in-between group differences (P = 0.4). However, a higher total urinary volume excretion in all alcohol compared with water interventions was observed (P = 0.01). Furthermore, plasma copeptin, sodium, and urinary osmolality levels increased significantly at the end of the observation period in all alcohol compared with water-only interventions (P = 0.02). In conclusion, initial copeptin suppression does not differ between alcohol or water interventions but seems to be prolonged in the alcohol interventions. This leads to increased volume loss followed by a counterregulation with increased copeptin levels and water retention after 720 min in alcohol compared with interventions. Additional sodium and/or water consumption with alcohol did not change the observed alcohol-induced effects.


Assuntos
Cerveja , Glicopeptídeos/metabolismo , Sódio/sangue , Equilíbrio Hidroeletrolítico , Água , Adulto , Consumo de Bebidas Alcoólicas , Ingestão de Líquidos , Glicopeptídeos/genética , Homeostase/efeitos dos fármacos , Humanos , Masculino , Adulto Jovem
10.
Mol Cell Proteomics ; 19(1): 11-30, 2020 01.
Artigo em Inglês | MEDLINE | ID: mdl-31591262

RESUMO

Glycosylation is a topic of intense current interest in the development of biopharmaceuticals because it is related to drug safety and efficacy. This work describes results of an interlaboratory study on the glycosylation of the Primary Sample (PS) of NISTmAb, a monoclonal antibody reference material. Seventy-six laboratories from industry, university, research, government, and hospital sectors in Europe, North America, Asia, and Australia submitted a total of 103 reports on glycan distributions. The principal objective of this study was to report and compare results for the full range of analytical methods presently used in the glycosylation analysis of mAbs. Therefore, participation was unrestricted, with laboratories choosing their own measurement techniques. Protein glycosylation was determined in various ways, including at the level of intact mAb, protein fragments, glycopeptides, or released glycans, using a wide variety of methods for derivatization, separation, identification, and quantification. Consequently, the diversity of results was enormous, with the number of glycan compositions identified by each laboratory ranging from 4 to 48. In total, one hundred sixteen glycan compositions were reported, of which 57 compositions could be assigned consensus abundance values. These consensus medians provide community-derived values for NISTmAb PS. Agreement with the consensus medians did not depend on the specific method or laboratory type. The study provides a view of the current state-of-the-art for biologic glycosylation measurement and suggests a clear need for harmonization of glycosylation analysis methods.


Assuntos
Anticorpos Monoclonais/química , Produtos Biológicos , Biofarmácia/métodos , Anticorpos Monoclonais/metabolismo , Glicômica/métodos , Glicopeptídeos/metabolismo , Glicosilação , Humanos , Laboratórios , Polissacarídeos/metabolismo , Processamento de Proteína Pós-Traducional , Proteômica/métodos
11.
J Endocrinol Invest ; 43(1): 21-30, 2020 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-31368050

RESUMO

COPEPTIN: Copeptin is secreted in equimolar amount to Arginine Vasopressin (AVP) but can easily be measured with a sandwich immunoassay. Both peptides, copeptin and AVP, show a high correlation. Accordingly, copeptin mirrors the amount of AVP in the circulation and its measurement provides an attractive marker in the differential diagnosis of diabetes insipidus. THE POLYURIA POLYDIPSIA SYNDROME: Diabetes insipidus-either central or nephrogenic-has to be differentiated from primary polydipsia. Differentiation is crucial since wrong treatment can have deleterious consequences. Since many decades, the "gold standard" for differential diagnosis has been the classical water deprivation test, which has several limitations leading to an overall limited diagnostic accuracy. In addition, the test has a long duration of 17 hours and is cumbersome for patients. Clinical signs and symptoms as well as MRI characteristics overlap between patients with diabetes insipidus and primary polydipsia. Direct measurement of AVP upon osmotic stimulation was first shown to overcome these limitations, but failed to enter clinical practice mainly due to technical limitations of the AVP assay. COPEPTIN AS DIAGNOSTIC TOOL IN THE POLYURIA POLYDIPSIA SYNDROME: We have recently shown that copeptin, without prior water deprivation, identifies patients with nephrogenic diabetes insipidus. On the other hand, for the more difficult differentiation between central diabetes insipidus and primary polydipsia, a copeptin level of 4.9 pmol/L stimulated with hypertonic saline infusion differentiates between these two entities with a high diagnostic accuracy, and is superior to the water deprivation test. It is important to note that close sodium monitoring during the hypertonic saline test is a prerequisite. CONCLUSION: Therefore, we propose that copeptin upon hypertonic saline infusion should become the new standard test in the differential diagnosis of diabetes insipidus.


Assuntos
Biomarcadores/metabolismo , Glicopeptídeos/metabolismo , Poliúria/classificação , Poliúria/diagnóstico , Diagnóstico Diferencial , Humanos , Poliúria/metabolismo
12.
Cancer Immunol Res ; 8(2): 167-178, 2020 02.
Artigo em Inglês | MEDLINE | ID: mdl-31831633

RESUMO

Patients with ulcerative colitis have an increased risk of developing colitis-associated colon cancer (CACC). Changes in glycosylation of the oncoprotein MUC1 commonly occur in chronic inflammation, including ulcerative colitis, and this abnormally glycosylated MUC1 promotes cancer development and progression. It is not known what causes changes in glycosylation of MUC1. Gene expression profiling of myeloid cells in inflamed and malignant colon tissues showed increased expression levels of inflammatory macrophage-associated cytokines compared with normal tissues. We analyzed the involvement of macrophage-associated cytokines in the induction of aberrant MUC1 glycoforms. A coculture system was used to examine the effects of M1 and M2 macrophages on glycosylation-related enzymes in colon cancer cells. M2-like macrophages induced the expression of the glycosyltransferase ST6GALNAC1, an enzyme that adds sialic acid to O-linked GalNAc residues, promoting the formation of tumor-associated sialyl-Tn (sTn) O-glycans. Immunostaining of ulcerative colitis and CACC tissue samples confirmed the elevated number of M2-like macrophages as well as high expression of ST6GALNAC1 and the altered MUC1-sTn glycoform on colon cells. Cytokine arrays and blocking antibody experiments indicated that the macrophage-dependent ST6GALNAC1 activation was mediated by IL13 and CCL17. We demonstrated that IL13 promoted phosphorylation of STAT6 to activate transcription of ST6GALNAC1. A computational model of signaling pathways was assembled and used to test IL13 inhibition as a possible therapy. Our findings revealed a novel cellular cross-talk between colon cells and macrophages within the inflamed and malignant colon that contributes to the pathogenesis of ulcerative colitis and CACC.See related Spotlight on p. 160.


Assuntos
Colite Ulcerativa/imunologia , Colite/complicações , Colo/imunologia , Neoplasias do Colo/imunologia , Glicopeptídeos/metabolismo , Células Mieloides/imunologia , Sialiltransferases/genética , Linhagem Celular Tumoral , Colite/imunologia , Colite Ulcerativa/metabolismo , Colite Ulcerativa/patologia , Colo/metabolismo , Neoplasias do Colo/metabolismo , Neoplasias do Colo/patologia , Biologia Computacional , Citocinas/genética , Citocinas/metabolismo , Glicosilação , Humanos , Inflamação/metabolismo , Interleucina-13/metabolismo , Ativação de Macrófagos/imunologia , Fator de Transcrição STAT6/metabolismo , Sialiltransferases/metabolismo , Transdução de Sinais
13.
Int J Mol Sci ; 22(1)2020 Dec 31.
Artigo em Inglês | MEDLINE | ID: mdl-33396442

RESUMO

We designed and synthesized amphiphilic glycopeptides with glucose or galactose at the C-terminals. We observed the protein-induced structural changes of the amphiphilic glycopeptide assembly in the lipid bilayer membrane using transmission electron microscopy (TEM) and Fourier transform infrared reflection-absorption spectra (FTIR-RAS) measurements. The glycopeptides re-arranged to form a bundle that acted as an ion channel due to the interaction among the target protein and the terminal sugar groups of the glycopeptides. The bundle in the lipid bilayer membrane was fixed on a gold-deposited quartz crystal microbalance (QCM) electrode by the membrane fusion method. The protein-induced re-arrangement of the terminal sugar groups formed a binding site that acted as a receptor, and the re-binding of the target protein to the binding site induced the closing of the channel. We monitored the detection of target proteins by the changes of the electrochemical properties of the membrane. The response current of the membrane induced by the target protein recognition was expressed by an equivalent circuit consisting of resistors and capacitors when a triangular voltage was applied. We used peanut lectin (PNA) and concanavalin A (ConA) as target proteins. The sensing membrane induced by PNA shows the specific response to PNA, and the ConA-induced membrane responded selectively to ConA. Furthermore, PNA-induced sensing membranes showed relatively low recognition ability for lectin from Ricinus Agglutinin (RCA120) and mushroom lectin (ABA), which have galactose binding sites. The protein-induced self-organization formed the spatial arrangement of the sugar chains specific to the binding site of the target protein. These findings demonstrate the possibility of fabricating a sensing device with multi-recognition ability that can recognize proteins even if the structure is unknown, by the protein-induced self-organization process.


Assuntos
Concanavalina A/química , Eletrodos , Glicopeptídeos/química , Bicamadas Lipídicas/química , Aglutinina de Amendoim/química , Lectinas de Plantas/química , Sítios de Ligação , Concanavalina A/metabolismo , Glicopeptídeos/metabolismo , Ouro , Canais Iônicos , Bicamadas Lipídicas/metabolismo , Aglutinina de Amendoim/metabolismo , Lectinas de Plantas/metabolismo
14.
J Biol Chem ; 294(49): 18769-18783, 2019 12 06.
Artigo em Inglês | MEDLINE | ID: mdl-31672921

RESUMO

Since the discovery of vancomycin in the 1950s, the glycopeptide antibiotics (GPAs) have been of great interest to the scientific community. These nonribosomally biosynthesized peptides are highly cross-linked, often glycosylated, and inhibit bacterial cell wall assembly by interfering with peptidoglycan synthesis. Interest in glycopeptide antibiotics covers many scientific disciplines, due to their challenging total syntheses, complex biosynthesis pathways, mechanism of action, and high potency. After intense efforts, early enthusiasm has given way to a recognition of the challenges in chemically synthesizing GPAs and of the effort needed to study and modify GPA-producing strains to prepare new GPAs to address the increasing threat of microbial antibiotic resistance. Although the preparation of GPAs, either by modifying the pendant groups such as saccharides or by functionalizing the N- or C-terminal moieties, is readily achievable, the peptide core of these molecules-the GPA aglycone-remains highly challenging to modify. This review aims to present a summary of the results of GPA modification obtained with the three major approaches developed to date: in vivo strain manipulation, total chemical synthesis, and chemoenzymatic synthesis methods.


Assuntos
Antibacterianos/química , Antibacterianos/metabolismo , Glicopeptídeos/química , Glicopeptídeos/metabolismo , Animais , Produtos Biológicos/química , Produtos Biológicos/metabolismo , Doenças Transmissíveis/metabolismo , Glicosilação , Humanos
15.
Bioconjug Chem ; 30(11): 2715-2726, 2019 11 20.
Artigo em Inglês | MEDLINE | ID: mdl-31580646

RESUMO

Glycosylation plays a myriad of roles in the immune system: Certain glycans can interact with specific immune receptors to kickstart a pro-inflammatory response, whereas other glycans can do precisely the opposite and ameliorate the immune response. Specific glycans and glycoforms can themselves become the targets of the adaptive immune system, leading to potent antiglycan responses that can lead to the killing of altered self- or pathogenic species. This hydra-like set of roles glycans play is of particular importance in cancer immunity, where it influences the anticancer immune response, likely playing pivotal roles in tumor survival or clearance. The complexity of carbohydrate biology requires synthetic access to glycoproteins and glycopeptides that harbor homogeneous glycans allowing the probing of these systems with high precision. One particular complicating factor in this is that these synthetic structures are required to be as close to the native structures as possible, as non-native linkages can themselves elicit immune responses. In this Review, we discuss examples and current strategies for the synthesis of natively linked single glycoforms of peptides and proteins that have enabled researchers to gain new insights into glycoimmunology, with a particular focus on the application of these reagents in cancer immunology.


Assuntos
Glicopeptídeos/imunologia , Glicoproteínas/imunologia , Doenças do Sistema Imunitário/patologia , Sistema Imunitário/imunologia , Sistema Imunitário/metabolismo , Polissacarídeos/imunologia , Animais , Glicopeptídeos/metabolismo , Glicoproteínas/metabolismo , Glicosilação , Humanos , Doenças do Sistema Imunitário/imunologia , Doenças do Sistema Imunitário/metabolismo , Polissacarídeos/metabolismo
16.
Andrologia ; 51(11): e13437, 2019 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-31637753

RESUMO

Pattern recognition receptors (PRRs) such as toll-like receptors (TLRs) and nuclear oligomerization domain (NOD) receptors along with antimicrobial proteins and peptides (AMPs) are crucial for innate immunity. The pathology of insulin-dependent diabetes mellitus is associated with the disrupted expression of TLRs, NODs and AMPs in the kidney, lungs and other organs. However, such a relation in the male reproductive tract is not yet investigated. In this study, we analysed the expression pattern of Tlr1-13, Nod1/2 receptors and AMPs (ß-defensins and defensin-like proteins of the Sperm-Associated Antigen 11 (Spag11) family) in the male reproductive tissues (caput, cauda and testis) obtained from diabetic or insulin-treated diabetic or untreated control rats. Alterations in the expression pattern of Tlr1-13, Nod 1/ 2, Defb1, 2, 21, 24, 27, 30 and Spag11a/ c/ t were observed under diabetic conditions. Administration of insulin to diabetic rats could modulate the expression pattern of only some these genes. Results of our study indicate perturbed gene expression profile of Tlrs, Nod1/2, Defbs and Spag11 isoforms in the epididymis and testis of diabetic rats, and this could be one of the important reasons for the increased risk of infections in the male genital tract.


Assuntos
Peptídeos Catiônicos Antimicrobianos/metabolismo , Defensinas/metabolismo , Diabetes Mellitus Experimental/metabolismo , Epididimo/metabolismo , Testículo/metabolismo , Receptores Toll-Like/metabolismo , Aloxano , Animais , Antígenos de Superfície/metabolismo , Diabetes Mellitus Experimental/tratamento farmacológico , Epididimo/efeitos dos fármacos , Glicopeptídeos/metabolismo , Insulina/farmacologia , Insulina/uso terapêutico , Masculino , Ratos Wistar , Testículo/efeitos dos fármacos , beta-Defensinas
17.
Rev Endocr Metab Disord ; 20(3): 283-294, 2019 09.
Artigo em Inglês | MEDLINE | ID: mdl-31656992

RESUMO

Arginine Vasopressin (AVP) and copeptin derive from the same precursor molecule. Due to the equimolar secretion, copeptin responds as rapidly as AVP to osmotic, hemodynamic and unspecific stress-related stimuli and both peptides show a very strong correlation. The physiological functions of AVP are homeostasis of fluid balance, vascular tonus and regulation of the endocrine stress response. In contrast, the exact function of copeptin remains unknown. Since copeptin, in contrast to AVP, can easily be measured with a sandwich immunoassay, its main function so far that it indirectly indicates the amount of AVP in the circulation. Copeptin has emerged as a useful measure in different diseases. On one hand, through its characteristics as a marker of stress, it provides a unique measure of the individual stress burden. As such, it is a prognostic marker in different acute diseases such as ischemic stroke or myocardial infarction. On the other side, it has emerged as a promising marker in the diagnosis of AVP-dependent fluid disorders. Copeptin reliably differentiates various entities of the polyuria polydipsia syndrome; baseline levels >20 pmol/L without prior fluid deprivation identify patients with nephrogenic diabetes insipidus, whereas levels measured upon osmotic stimulation with hypertonic saline or upon non-osmotic stimulation with arginine differentiate primary polydipsia from central diabetes insipidus. In patients with hyponatremia, low levels of copeptin together with low urine osmolality identify patients with primary polydipsia, but copeptin levels overlap in all other causes of hyponatremia, limiting its diagnostic use in hyponatremia. Copeptin has also been put forward as predictive marker for autosomal dominant polycystic kidney disease and for diabetes mellitus, but more studies are needed to confirm these findings.


Assuntos
Arginina Vasopressina/metabolismo , Diabetes Insípido/metabolismo , Glicopeptídeos/metabolismo , Vasopressinas/metabolismo , Feminino , Humanos , Masculino
18.
Int J Mol Sci ; 20(19)2019 Oct 01.
Artigo em Inglês | MEDLINE | ID: mdl-31581426

RESUMO

The advent of multidrug resistance among pathogenic bacteria has attracted great attention worldwide. As a response to this growing challenge, diverse studies have focused on the development of novel anti-infective therapies, including antimicrobial peptides (AMPs). The biological properties of this class of antimicrobials have been thoroughly investigated, and membranolytic activities are the most reported mechanisms by which AMPs kill bacteria. Nevertheless, an increasing number of works have pointed to a different direction, in which AMPs are seen to be capable of displaying non-lytic modes of action by internalizing bacterial cells. In this context, this review focused on the description of the in vitro and in vivo antibacterial and antibiofilm activities of non-lytic AMPs, including indolicidin, buforin II PR-39, bactenecins, apidaecin, and drosocin, also shedding light on how AMPs interact with and further translocate through bacterial membranes to act on intracellular targets, including DNA, RNA, cell wall and protein synthesis.


Assuntos
Antibacterianos/farmacologia , Peptídeos Catiônicos Antimicrobianos/farmacologia , Bactérias/efeitos dos fármacos , Membrana Celular/efeitos dos fármacos , Membrana Celular/metabolismo , Animais , Peptídeos Catiônicos Antimicrobianos/metabolismo , Bactérias/metabolismo , Glicopeptídeos/metabolismo , Insetos , Biossíntese de Proteínas
19.
Microb Cell Fact ; 18(1): 182, 2019 Oct 26.
Artigo em Inglês | MEDLINE | ID: mdl-31655587

RESUMO

BACKGROUND: Oxidative stress caused by inevitable hostile conditions during fermentative process was the most serious threat to the survival of the well-known industrial microorganism Corynebacterium glutamicum. To survive, C. glutamicum developed several antioxidant defenses including millimolar concentrations of mycothiol (MSH) and protective enzymes. Glutathione (GSH) S-transferases (GSTs) with essentially defensive role in oxidative stress have been well defined in numerous microorganisms, while their physiological and biochemical functions remained elusive in C. glutamicum thus far. RESULTS: In the present study, we described protein NCgl1216 belonging to a novel MSH S-transferase Xi class (MstX), considered as the equivalent of GST Xi class (GSTX). MstX had a characteristic conserved catalytic motif (Cys-Pro-Trp-Ala, C-P-W-A). MstX was active as thiol transferase, dehydroascorbate reductase, mycothiolyl-hydroquinone reductase and MSH peroxidase, while it showed null activity toward canonical GSTs substrate as 1-chloro-2,4-dinitrobenzene (CDNB) and GST Omega's specific substance glutathionyl-acetophenones, indicating MstX had some biochemical characteristics related with mycoredoxin (Mrx). Site-directed mutagenesis showed that, among the two cysteine residues of the molecule, only the residue at position 67 was required for the activity. Moreover, the residues adjacent to the active Cys67 were also important for activity. These results indicated that the thiol transferase of MstX operated through a monothiol mechanism. In addition, we found MstX played important role in various stress resistance. The lack of C. glutamicum mstX gene resulted in significant growth inhibition and increased sensitivity under adverse stress condition. The mstX expression was induced by stress. CONCLUSION: Corynebacterium glutamicum MstX might be critically involved in response to oxidative conditions, thereby giving new insight in how C. glutamicum survived oxidative stressful conditions.


Assuntos
Proteínas de Bactérias/química , Corynebacterium glutamicum/metabolismo , Cisteína/metabolismo , Glutationa Transferase/química , Glicopeptídeos/metabolismo , Inositol/metabolismo , Oxirredução , Estresse Oxidativo
20.
J Mater Chem B ; 7(37): 5695-5703, 2019 09 25.
Artigo em Inglês | MEDLINE | ID: mdl-31482162

RESUMO

The long-term cryopreservation of cells, tissues and organs is limited by both osmotic stress and ice injury during freezing or thawing. Introduction of biocompatible cryoprotective agents is promising for cell cryopreservation based on membrane stabilization of glycopeptides and trehalose. Herein, a series of trehalose-functional (glyco)peptides were synthesized by successively tethering Nω-(4-toluenesulfonyl)-l-arginine and carboxylated trehalose to the side amino groups of ε-polylysine. The specifically synthesized glycopeptide demonstrated enhanced cryosurvival of sheep red blood cells (RBCs) from 49.1 ± 0.9% to 75.0 ± 2.4% at pH 7.4, along with trehalose molecules. Cryopreservation via membrane stabilization was proposed such that the glycopeptide could attach on the cell membrane surface of RBCs to protect the cell membrane from osmotic injury by electrostatic interactions, π-π stacking and hydrogen bonding. Meanwhile, trehalose could prevent RBCs from ice injury via ice recrystallization inhibition activity. Thus, membrane protection for preventing osmotic stress and ice injury could be boosted via synergistic interaction between the glycopeptide and trehalose. The combined usage of biocompatible glycopeptides and trehalose could potentially be applied in the glycerol-free cryopreservation of cells and tissues in biomedicine.


Assuntos
Criopreservação/métodos , Eritrócitos/metabolismo , Glicopeptídeos/metabolismo , Trealose/metabolismo , Eritrócitos/citologia
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