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1.
The majority of synaptic plasma membrane components are glycosylated. It is now widely accepted that this post-translational modification is crucial during the establishment, maintenance and function of the nervous system. Despite its significance, structural information about the glycosylation of nervous system specific glycoproteins is very limited. In the present study the major glycan structures of the chicken synaptic plasma membrane (SPM) associated glycoprotein glycans were determined. N-glycans were released by hydrazinolysis, labelled with 2-aminobenzamide, treated with neuraminidase and subsequently fractionated by size exclusion chromatography. Individual fractions were characterized by the combination of high-pressure liquid chromatography, exoglycosidase treatment or reagent array analysis method (RAAM). In addition to oligomannose-type glycans, core-fucosylated complex glycans with biantennary bisecting glycans carrying the LewisX epitope were most abundant. The overall chicken glycan profile was strikingly similar to the rat brain glycan profile. The presence of the LewisX determinant in relatively large proportions suggests a tissue-specific function for these glycans.  相似文献   

2.
Over a half of all proteins are glycosylated, and their proper glycosylation is essential for normal function. Unfortunately, because of structural complexity of nonlinear branched glycans and the absence of genetic template for their synthesis, the knowledge about glycans is lagging significantly behind the knowledge about proteins or DNA. Using a recently developed quantitative high throughput glycan analysis method we quantified components of the plasma N-glycome in 99 children with attention-deficit hyperactivity disorder (ADHD), 81 child and 5 adults with autism spectrum disorder, and a total of 340 matching healthy controls. No changes in plasma glycome were found to associate with autism spectrum disorder, but several highly significant associations were observed with ADHD. Further structural analysis of plasma glycans revealed that ADHD is associated with increased antennary fucosylation of biantennary glycans and decreased levels of some complex glycans with three or four antennas. The design of this study prevented any functional conclusions about the observed associations, but specific differences in glycosylation appears to be strongly associated with ADHD and warrants further studies in this direction.  相似文献   

3.
Characterization of the N-glycans from human pancreatic ribonuclease (RNase 1) isolated from healthy pancreas and from pancreatic adenocarcinoma tumor cells (Capan-1 and MDAPanc-3) revealed completely different glycosylation patterns. RNase 1 from healthy cells contained neutral complex biantennary structures, with smaller amounts of tri- and tetraantennary compounds, and glycans with poly-N-acetyllactosamine extensions, all extensively fucosylated. In contrast, RNase 1 glycans from tumor cells (Capan-1) were fucosylated hybrid and complex biantennary glycans with GalNAc-GlcNAc antennae. RNase 1 glycans from Capan-1 and MDAPanc-3 cells also contained sialylated structures completely absent in the healthy pancreas. Some of these features provide distinct epitopes that were clearly detected using monoclonal antibodies against carbohydrate antigens. Thus monoclonal antibodies to Lewis(y) reacted only with normal pancreatic RNase 1, whereas, in contrast, monoclonal antibodies to sialyl-Lewis(x) and sialyl-Lewis(a) reacted only with RNase 1 secreted from the tumor cells. These glycosylation changes in a tumor-secreted protein, which reflect fundamental changes in the enzymes involved in the glycosylation pathway, open up the possibility of using serum RNase 1 as a tumor marker of pancreatic adenocarcinoma.  相似文献   

4.
Variable (Fv) domain N-glycosylation sites are found in approximately 20% of human immunoglobulin Gs (IgGs) in addition to the conserved N-glycosylation sites in the C(H)2 domains. The carbohydrate structures of the Fv glycans and their impact on in vivo half-life are not well characterized. Oligosaccharide structures in a humanized anti-Abeta IgG1 monoclonal antibody (Mab) with an N-glycosylation site in the complementary determining region (CDR2) of the heavy chain variable region were elucidated by LC/MS analysis following sequential exoglycosidase treatments of the endoproteinase Lys-C digest. Results showed that the major N-linked oligosaccharide structures in the Fv region have three characteristics (core-fucosylated biantennary oligosaccharides with one or two N-glycolylneuraminic acid [NeuGc] residues, zero or one alpha-linked Gal residue, and zero or one beta-linked GalNAc residue), whereas N-linked oligosaccharides in the Fc region contained typical Fc glycans (core-fucosylated, biantennary oligosaccharides with zero to two Gal residues). To elucidate the contribution of Fv glycans to the half-life of the antibody, a method that allows capture of the Mab and determination of its glycan structures at various time points after administration to mice was developed. Anti-Abeta antibody in mouse serum was immunocaptured by immobilized goat anti-human immunoglobulin Fc(gamma) antibody resin, and the captured material was treated with papain to generate Fab and Fc for LC/MS analysis. Different glycans in the Fc region showed the same clearance rate as demonstrated previously. In contrast to many other non-antibody glycosylated therapeutics, there is no strong correlation between oligosaccharide structures in the Fv region and their clearance rates in vivo. Our data indicated that biantennary oligosaccharides lacking galactosylation had slightly faster clearance rates than other structures in the Fv domain.  相似文献   

5.
R24 is a mouse IgG3 monoclonal antibody (mab) that reacts with the ganglioside GD3 expressed by cells of neuroectodermal origin. The anti-tumor activity of R24 has been demonstrated in initial phase I and pilot trials in patients suffering from metastatic melanoma. The purpose of this study was to investigate the biotechnological production and particularly the glycosylation of this clinically important antibody. Growth, metabolism, and IgG production of R24 secreting hybridoma cells were analyzed on 1 L bioreactor bench scale using repeated-batch mode. The amount of 57 mg of pure mab was obtained from 1.6 L crude supernatant by protein A chromatography. Western blot binding assays with sugar-specific lectins revealed glycosylation of the heavy chains, whereas no carbohydrates were detectable on the light chains. Because glycosylation is essential for antibody effector functions in vivo (such as complement fixation or binding to macrophage Fc receptors), mab R24 was subjected to both enzymatic deglycosylation using PNGase F and chemical deglycosylation by hydrazinolysis. Released glycans were structurally characterized by high pH anion exchange chromatography with pulsed amperometric detection (HPAEC-PAD), matrix assisted laser desorption ionization time-of-flight (MALDI-TOF), and electrospray ionization quadrupole time-of-flight (ESI-QTOF) mass spectrometry. Six major biantennary chains of the complex glycosylation phenotype were found with variations in galactosylation and core fucosylation. The predominant N-linked structure, indicating the high degree of agalactosyl glycoforms, was the agalacto biantennary chain with a relative percentage of 57% (51% core-fucosylated, 6% nonfucosylated). The second most abundant oligosaccharide was the monogalacto biantennary chain amounting to 30% (26% core- and 4% nonfucosylated). The antibody contained 0.46 microg sialic acid per mg protein, which splits into 0.243 microg Neu5Gc and 0.217 microg Neu5Ac, corresponding to a Neu5Ac:Neu5Gc ratio of 1:1.06. Furthermore, the antigen specificity of R24 was determined by immunodetection of GD3 on thin-layer chromatograms, and real time GD3-antibody binding interactions were measured with an optical biosensor (BIAcore). From the structural data obtained in this study it is concluded that glycosylation of the antibody may be important in the clinical outcome of targeted anti-cancer immunotherapy.  相似文献   

6.
Despite the importance of protein glycosylation in all physiological and pathological processes and their potential as diagnostic markers and drug targets, the glycome of children is still unexplored. We analyzed N-linked plasma and IgG glycomes in 170 children and adolescents between 6 and 18 years of age. The results showed large biological variability at the population level as well as a large number of associations between different glycans and age. The plasma N-glycome of younger children was found to contain a larger proportion of large complex glycan structures (r = -0.71 for tetrasialylated glycans; r = -0.41 for trisialylated glycans) as well as an increase in disialylated biantennary structures (r = 0.55) with age. Core fucosylation and the level of agalactosylated plasma and IgG glycans decreased while digalactosylated glycans increased with age. This pattern of age-dependent changes in children differs from changes reported in adult population in both, direction and the intensity of changes. Also, sex differences are much smaller in children than in adults and are present mainly during puberty. These important observations should be accounted for when glycan-based diagnostic tests or therapeutics are being developed or evaluated.  相似文献   

7.
Gelatinase B is a matrix metalloproteinase (MMP-9) involved in tissue remodeling, development, cancer, and inflammation. Neutrophils produce three major forms of (pro)gelatinase B: 92 kDa monomers, homodimers, and complexes of gelatinase B covalently bound to neutrophil gelatinase B-associated lipocalin (NGAL). In contrast to the case for other proteinases, little information about the glycosylation of any natural human MMP is available. Here, both gelatinase B and NGAL were purified from human peripheral blood neutrophils, and the entire contents of the released N- and O-glycan pools were analyzed simultaneously using recently developed high-performance liquid chromatography-based technology. The results are discussed within the context of the domain structure of gelatinase B and a molecular model of NGAL based on data from this study and the three-dimensional nuclear magnetic resonance (NMR) structure of the protein. More than 95% of the N-linked glycans attached to both gelatinase B and NGAL were partially sialylated, core-fucosylated biantennary structures with and without outer arm fucose. The O-linked glycans, which were estimated to comprise approximately 85% of the total sugars on gelatinase B, mainly consisted of type 2 cores with Galbeta1,4GlcNAc (lactosamine) extensions, with or without sialic acid or outer arm fucose. This paper also contains the first report of O-linked glycans attached to NGAL. Although both proteins were isolated from neutrophils and contained O-linked glycans mainly with type 2 cores, the glycans attached to individual serine/threonine residue(s) in NGAL were significantly smaller than those on gelatinase B. In contrast to NGAL, gelatinase B contains a region rich in Ser, Thr, and Pro typical of O-glycosylated mucin-like domains.  相似文献   

8.
Defective galactosylation of serum transferrin in galactosemia   总被引:6,自引:1,他引:5  
The glycosylation of serum transferrin from galactosemic patients with a deficiency of galactose-1-phosphate uridyl transferase (EC 2. 7.7 12) is abnormal but becomes normal after treatment with a galactose-free diet. To understand the structural and biochemical basis of the abnormal glycosylation, transferrin was purified from the serum of untreated and treated galactosemic patients and normal controls and the N-linked glycans analyzed by HPLC. The glycans from normal transferrin consisted predominantly (86%) of the disialylated biantennary complex type. The glycans from untreated galactosemic patients were more heterogeneous and contained four major truncated glycans in addition to a smaller amount (13%) of the disialylated biantennary complex type. The truncated glycans were deficient in galactose and sialic acid and their structures were consistent with a decrease in galactosyltransferase activity in hepatocytes, the probable cells of origin of the transferrin. This is postulated to be due to direct inhibition of the galactosyltransferase activity by the accumulated galactose-1-phosphate or to an effect on the formation of UDP- galactose, the donor substrate in the reaction. After treatment the proportion of the truncated glycans decreased and the proportion of the disialylated biantennary complex type increased, returning almost but never completely to normal, even after prolonged treatment in some cases. There was no clear relationship between the length of treatment and the normalization of glycosylation and the level of galactose-1- phosphate in red blood cells, the usual parameter for monitoring the treatment of galactosemics. It is suggested that the persistence of abnormally glycosylated proteins may contribute to the long-term complications in galactosemia.   相似文献   

9.
A series of enzymatic substitutions modifies the basic structure of complex-type biantennary N-glycans. Among them, a beta1,2-linked N-acetylglucosamine residue is introduced to the central mannose moiety of the core-fucosylated oligosaccharide by N-acetylglucosaminyltransferase VII. This so-called LEC14 epitope can undergo galactosylation at the beta1,2-linked N-acetylglucosamine residue. Guided by the hypothesis that structural modifications in the N-glycan alter its capacity to serve as ligand for lectins, we prepared a neoglycoprotein with the extended LEC14 N-glycan and tested its properties in three different assays. In order to allow comparison to previous results on other types of biantennary N-glycans the functionalization of the glycans for coupling and assay conditions were deliberately kept constant. Compared to the core-fucosylated N-glycan no significant change in affinity was seen when testing three galactoside-specific proteins. However, cell positivity in flow cytofluorimetry was enhanced in six of eight human tumor lines. Analysis of biodistribution in tumor-bearing mice revealed an increase of blood clearance by about 40%, yielding a favorable tumor/blood ratio. Thus, the extended LEC14 motif affects binding properties to cellular lectins on cell surfaces and organs when compared to the core-fucosylated biantennary N-glycan. The results argue in favor of the concept of viewing substitutions as molecular switches for lectin-binding affinity. Moreover, they have potential relevance for glycoengineering of reagents in tumor imaging.  相似文献   

10.
The N-linked glycosylation of recombinant human CD59, expressed in Chinese hamster ovary (CHO) cells with and without a membrane anchor, was compared to examine the effect of the anchor on glycan processing. N-Linked glycans were released with peptide-N-glycosidase F (PNGase F) within gel from SDS-PAGE-isolated soluble and glycosylphosphatidylinositol (GPI)-anchored human CD59 expressed in CHO cells. The anchored form contained core-fucosylated neutral and sialylated bi-, tri-, and tetraantennary glycans with up to four N-acetyllactosamine extensions. Exoglycosidase digestions and analysis by matrix-assisted laser desorption/ionization (MALDI) mass spectrometry were used to define the relative amounts of the bi-, tri-, and tetraantennary glycans and to investigate the distribution of N-acetyllactosamine extensions between their antennae. Biantennary structures accounted for about 60% of the glycans, 30% of the triantennary structures, and about 10% of the tetraantennary structures. For tri- and tetraantennary glycans, those with extended antennae were found to be more abundant than those without extensions. The soluble form of CD59, expressed in CHO cells without the GPI anchor signal sequence, consisted almost entirely (97%) of biantennary glycans, of which 81% were unmodified, 17% contained one N-acetyllactosamine extension, and 2% contained two extensions. No compounds with longer extensions were found. A MALDI spectrum of the intact glycoprotein showed a distribution of glycans that matched those released with PNGase F. In addition, the protein was substituted with several small glycans, such as HexNAc, HexNAc-->Fuc, and HexNAc-->HexNAc, probably as the result of degradation of the mature N-linked glycans. The results show that the presence of the anchor increases the extent of glycan processing, possibly as the result of longer exposure to the glycosyltransferases or to a closer proximity of the protein to these enzymes.  相似文献   

11.
All four subclasses of human serum IgG contain a single N-glycosylation site in the constant region of their heavy chain, which is occupied by biantennary, largely core-fucosylated and partially truncated oligosaccharides, that may carry a bisecting N-acetylglucosamine and sialic acid residues. IgG glycosylation has been shown to be altered under various physiological and pathological circumstances. IgG N-glycan profiles vary with age, and galactosylation for example is enhanced during pregnancy. Several diseases including rheumatoid arthritis are associated with a reduction in galactosylation of the IgG N-glycans. Here, we describe a robust method for the isolation of IgG subclasses using protein A (binds IgG1, IgG2, and IgG4) and protein G (binds additionally IgG3) at the 96-well plate level, which is suitable for automation. Isolated IgGs were digested with trypsin, and obtained glycopeptides were analyzed by nano-LC-MS. Glycopeptides were characterized by CID as well as electron transfer dissociation (ETD). The method provided glycosylation profiles for IgG1, IgG2, IgG3, and IgG4 and revealed distinct differences in N-glycosylation between the four IgG subclasses. The changes in galactosylation associated with rheumatoid arthritis could readily be monitored. This method is suitable for the subclass-specific analysis of IgG glycosylation from clinical samples.  相似文献   

12.
Ovarian carcinoma is the leading cause of death from gynecological cancers in many Western countries. Aberrant glycosylation is an important aspect in malignant transformation and consequently in ovarian cancer. In this study, a detailed structure analysis of the N-linked glycans from total glycoproteins from the SKOV3 ovarian carcinoma cell line and from a recombinantly expressed secretory glycoprotein, erythropoietin (EPO), produced from the same cells has been performed using high-performance anion exchange chromatography with pulsed amperometric detection and matrix-assisted laser desorption/ionization time-of-flight mass spectrometry. Total cellular N-glycans contained high-mannose type and proximally fucosylated complex type partially agalactosylated structures. On the other hand, the recombinant human EPO secreted from SKOV3 cells contained predominantly core-fucosylated tetraantennary structures, which were partially lacking one or two galactose residues, and partially contained the LacdiNAc motif. Only minor amounts of di- and triantennary complex-type glycans were found, and high-mannose-type glycans were not present in the secreted EPO protein. A large amount of N-acetylneuraminic acid in α2,3-linkage was detected as well. Endogenous glycoproteins were also found to contain the LacdiNAc motif in N-linked glycans. This work contributes to the knowledge of the glycosylation of a human ovarian cancer cell line. It also establishes the basis to further explore high-mannose-type glycans, and the LacdiNAc motif as possible markers of ovarian carcinoma.  相似文献   

13.
Haptoglobin is one of acute phase glycoproteins often used as markers in glycopathology studies. In this work the oligosaccharide structures of haptoglobin from 'healthy' subjects have been studied in detail, taking into consideration the possible dependence of glycosylation on the phenotype. About 75% of charged haptoglobin glycans were of biantennary complex structure, and some of them lacked one terminal sialic acid molecule. Triantennary structures made up almost 25% of the charged glycans pool, and highly branched tetrasialylated oligosaccharides did not exceed 1%. The main difference between haptoglobin derived from the sample of pooled 44 sera and from the 2-2 phenotype individual concerned the relative content of trisialylated oligosaccharide with one 2-3 linked sialic acid residue. The oligosaccharide profile of haptoglobin derived from serum of a patient suffering from congenital disorder of glycosylation was compared to 'healthy' controls. It was shown, that four main glycans are identical in patient and 'normal' haptoglobins. Some alterations were found in the relative content of mono-, bi-, and trisialylated glycans as well as in the appearance of some tracely abundant oligosaccharides in haptoglobin of the patient with congenital disorder of glycosylation.  相似文献   

14.
The N-linked oligosaccharides were released from the phospholipase A2 (PLA) with glycopeptidases and reductively aminated with the chromophore, p-aminobenzoic acid ethyl ester (ABEE). The ABEE-labeled oligosaccharides were separated by microbore high-performance liquid chromatography (micro-HPLC) using a reversed-phase column and analyzed by electrospray mass spectrometry. Differentiation between alpha-1,3 and alpha-1,6 core-fucosylated glycans was achieved by comparison the glycans released by glycopeptidases peptide-N-glycanase A (PNase A) and peptide-N-glycanase F (PNase F). All N-linked oligosaccharides except 3B and 3C could be identified in this approach. The analysis of PLA oligosaccharides from the venom of individual bees indicated that glycosylation patterns between the younger and the older bees were similar.  相似文献   

15.
The extracellular matrix (ECM) molecules play important roles in many biological and pathological processes. During tissue remodeling, the ECM molecules that are glycosylated are different from those of normal tissue owing to changes in the expression of many proteins that are responsible for glycan synthesis. Vitronectin (VN) is a major ECM molecule that recognizes integrin on hepatic stellate cells (HSCs). The present study attempted to elucidate how changes in VN glycans modulate the survival of HSCs, which play a critical role in liver regeneration. Plasma VN was purified from partially hepatectomized (PH) and sham-operated (SH) rats at 24 h after operation and non-operated (NO) rats. Adhesion of rat HSCs (rHSCs), together with phosphorylation of focal adhesion kinase, in PH-VN was decreased to one-half of that in NO- or SH-VN. Spreading of rHSCs on desialylated NO-VN was decreased to one-half of that of control VN, indicating the importance of sialylation of VN for activation of HSCs. Liquid chromatography/multiple-stage mass spectrometry analysis of Glu-C glycopeptides of each VN determined the site-specific glycosylation. In addition to the major biantennary complex-type N-glycans, hybrid-type N-glycans were site-specifically present at Asn167. Highly sialylated O-glycans were found to be present in the Thr110–Thr124 region. In PH-VN, the disialyl O-glycans and complex-type N-glycans were decreased while core-fucosylated N-glycans were increased. In addition, immunodetection after two-dimensional PAGE indicated the presence of hyper- and hyposialylated molecules in each VN and showed that hypersialylation was markedly attenuated in PH-VN. This study proposes that the alteration of VN glycosylation modulates the substrate adhesion to rat HSCs, which is responsible for matrix restructuring.  相似文献   

16.
N-acetylglucosaminyltransferase V (GnT-V) catalyzes the addition of a beta1,6-linked GlcNAc to the alpha1,6 mannose of the trimannosyl core to form tri- and tetraantennary N-glycans and contains six putative N-linked sites. We used mass spectrometry techniques combined with exoglycosidase digestions of recombinant human GnT-V expressed in CHO cells, to identify its N-glycan structures and their sites of expression. Release of N-glycans by PNGase F treatment, followed by analysis of the permethylated glycans using MALDI-TOF MS, indicated a range of complex glycans from bi- to tetraantennary species. Mapping of the glycosylation sites was performed by enriching for trypsin-digested glycopeptides, followed by analysis of each fraction with Q-TOF MS. Predicted tryptic glycopeptides were identified by comparisons of theoretical masses of peptides with various glycan masses to the masses of the glycopeptides determined experimentally. Of the three putative glycosylation sites in the catalytic region, peptides containing sites Asn 334, 433, and 447 were identified as being N-glycosylated. Asn 334 is glycosylated with only a biantennary structure with one or two terminating sialic acids. Sites Asn 433 and 447 both contain structures that range from biantennary with two sialic acids to tetraantennary terminating with four sialic acids. The predominant glycan species found on both of these sites is a triantennary with three sialic acids. The appearance of only biantennary glycans at site Asn 433, coupled with the appearance of more highly branched structures at Asn 334 and 447, demonstrates that biantennary acceptors present at different sites on the same protein during biosynthesis can differ in their accessibility for branching by GnT-V.  相似文献   

17.
Hamster sarcoma virus (HSV) transformation of Nil-8 fibroblasts is associated with an increase in the average size of N-acetyllactosamine (complex) type N-linked glycans due to an increase in both the average number of branches/chain and in the fraction of N-linked glycans containing poly(GlcNAc(beta 1,3) Gal-(beta 1,4)) (polylactosaminylglycan) chains. Analysis of glycopeptides from the envelope glycoproteins of Sindbis virus and vesicular stomatitis virus (VSV) grown in Nil-8 and Nil/HSV cells indicated that the transformation-associated shift to larger N-linked oligosaccharides selectively affects some glycosylation sites far more than others. Glycosylation of the Sindbis virus glycoproteins and of Asn-179 of VSV G was similar in Nil-8 and Nil/HSV cells; oligosaccharide processing generally did not proceed beyond the biantennary complex stage. In contrast, Asn-336 of VSV G carried primarily biantennary complex glycans in Nil-8-grown virus (ratio, triantennary, and larger to biantennary complex glycans (tri+/bi) = 0.5) but more highly branched structures in Nil/HSV-grown virus (tri+/bi = 8.1). All of the triantennary or larger oligosaccharides from Asn-336 of Nil/HSV-grown VSV G bound to leukoagglutinating phytohemagglutinin-agarose, indicating the presence of a branch attached to the Man3GlcNAc2 core via a beta 1,6-linked GlcNAc residue and suggesting that increased UDP-GlcNAc:alpha-D-mannoside beta 1,6-N-acetylglucosaminyl transferase V (GlcNAc transferase V) activity accompanied transformation. At least 20% of these leukoagglutinating phytohemagglutinin-binding oligosaccharides were sensitive to an enzyme specific for polylactosaminylglycan chains, Escherichia freundii endo-beta-galactosidase.  相似文献   

18.
Recently, our group reported the expression of recombinant human erythropoietin in goat milk (rhEPO-milk) as well as in the mammary epithelial cell line GMGE (EPO-GMGE) by cell culture using the adenoviral transduction system. N-Glycosylation characterization of rhEPO-milk by Normal-Phase HPLC profiling of the fluorophore, 4-aminobenzoic acid-labeled enzymatically released N-glycan pool from rhEPO-goat milk, combined with MALDI, ESI-MS and LC/MS, revealed that low branched, core-fucosylated, N-glycans predominate. The labeled N-glycans were separated into neutral and charged fractions by anion exchange chromatography and the charged N-glycans were found to be mostly α2,6-monosialylated with Neu5Ac or Neu5Gc in a ratio of 1:1. Unlike the N-glycans from rhEPO produced in CHO cells, where the glycans are multiantennary highly sialylated, core-fucosylated oligosaccahrides, or even in the goat mammary gland epithelial cell line cultured in vitro in which multiantennary, core- and outer-arm fucosylated, monosialylated N-glycans are the most abundant species, a large proportion of the N-glycans from rhEPO-milk were monosialylated, biantennary, antennae mostly terminating with the more unusual GalNAc-GlcNAc motive and without outer-arm fucosylation. These findings, emphasizing the difference in the N-glycan repertoire between the rhEPO-milk and EPO-GMGE, are consistent with the principle that glycosylation is cell-type dependent and that the cell environment is crucial as well.  相似文献   

19.
Lung cancer has a poor prognosis and a 5-year survival rate of 15%. Therefore, early detection is vital. Diagnostic testing of serum for cancer-associated biomarkers is a noninvasive detection method. Glycosylation is the most frequent post-translational modification of proteins and it has been shown to be altered in cancer. In this paper, high-throughput HILIC technology was applied to serum samples from 100 lung cancer patients, alongside 84 age-matched controls and significant alterations in N-linked glycosylation were identified. Increases were detected in glycans containing Sialyl Lewis X, monoantennary glycans, highly sialylated glycans and decreases were observed in core-fucosylated biantennary glycans, with some being detectable as early as in Stage I. The N-linked glycan profile of haptoglobin demonstrated similar alterations to those elucidated in the total serum glycome. The most significantly altered HILIC peak in lung cancer samples includes predominantly disialylated and tri- and tetra-antennary glycans. This potential disease marker is significantly increased across all disease groups compared to controls and a strong disease effect is visible even after the effect of smoking is accounted for. The combination of all glyco-biomarkers had the highest sensitivity and specificity. This study identifies candidates for further study as potential biomarkers for the disease.  相似文献   

20.
Differentiation and activation of lymphocytes are documented to result in changes in glycosylation associated with biologically important consequences. In this report, we have systematically examined global changes in N-linked glycosylation following activation of murine CD4 T cells, CD8 T cells, and B cells by MALDI-TOF mass spectrometry profiling, and investigated the molecular basis for those changes by assessing alterations in the expression of glycan transferase genes. Surprisingly, the major change observed in activated CD4 and CD8 T cells was a dramatic reduction of sialylated biantennary N-glycans carrying the terminal NeuGcalpha2-6Gal sequence, and a corresponding increase in glycans carrying the Galalpha1-3Gal sequence. This change was accounted for by a decrease in the expression of the sialyltransferase ST6Gal I, and an increase in the expression of the galactosyltransferase, alpha1-3GalT. Conversely, in B cells no change in terminal sialylation of N-linked glycans was evident, and the expression of the same two glycosyltransferases was increased and decreased, respectively. The results have implications for differential recognition of activated and unactivated T cells by dendritic cells and B cells expressing glycan-binding proteins that recognize terminal sequences of N-linked glycans.  相似文献   

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