scholarly journals Tissue-specific N-glycosylation, site-specific oligosaccharide patterns and lentil lectin recognition of rat Thy-1.

1987 ◽  
Vol 6 (5) ◽  
pp. 1233-1244 ◽  
Author(s):  
R. B. Parekh ◽  
A. G. Tse ◽  
R. A. Dwek ◽  
A. F. Williams ◽  
T. W. Rademacher
Keyword(s):  
Glycosylation Site ◽  
Site Specific ◽  
Tissue Specific ◽  
Lentil Lectin ◽  
Lectin Recognition

scholarly journals Oxonium Ion Guided Analysis of Quantitative Proteomics Data Reveals Site-Specific O-Glycosylation of Anterior Gradient Protein 2 (AGR2)

2021 ◽  
Vol 22 (10) ◽  
pp. 5369
Author(s):  
Martina Pirro ◽  
Yassene Mohammed ◽  
Arnoud H. de Ru ◽  
George M. C. Janssen ◽  
Rayman T. N. Tjokrodirijo ◽  
...  
Keyword(s):  
Cell Lines ◽  
Quantitative Proteomics ◽  
Glycosylation Site ◽  
Total Cell ◽  
Tandem Mass ◽  
Proteomics Data ◽  
Site Specific ◽  
Oxonium Ions ◽  
Tandem Mass Tag ◽  
Colorectal Cancer Cell Lines

Developments in mass spectrometry (MS)-based analyses of glycoproteins have been important to study changes in glycosylation related to disease. Recently, the characteristic pattern of oxonium ions in glycopeptide fragmentation spectra had been used to assign different sets of glycopeptides. In particular, this was helpful to discriminate between O-GalNAc and O-GlcNAc. Here, we thought to investigate how such information can be used to examine quantitative proteomics data. For this purpose, we used tandem mass tag (TMT)-labeled samples from total cell lysates and secreted proteins from three different colorectal cancer cell lines. Following automated glycopeptide assignment (Byonic) and evaluation of the presence and relative intensity of oxonium ions, we observed that, in particular, the ratio of the ions at m/z 144.066 and 138.055, respectively, could be used to discriminate between O-GlcNAcylated and O-GalNAcylated peptides, with concomitant relative quantification between the different cell lines. Among the O-GalNAcylated proteins, we also observed anterior gradient protein 2 (AGR2), a protein which glycosylation site and status was hitherto not well documented. Using a combination of multiple fragmentation methods, we then not only assigned the site of modification, but also showed different glycosylation between intracellular (ER-resident) and secreted AGR2. Overall, our study shows the potential of broad application of the use of the relative intensities of oxonium ions for the confident assignment of glycopeptides, even in complex proteomics datasets.

scholarly journals Are we there yet? A machine learning architecture to predict organotropic metastases

2021 ◽  
Vol 14 (1) ◽  
Author(s):  
Michael Skaro ◽  
Marcus Hill ◽  
Yi Zhou ◽  
Shannon Quinn ◽  
Melissa B. Davis ◽  
...  
Keyword(s):  
Machine Learning ◽  
Cancer Metastasis ◽  
Driving Forces ◽  
The Cancer Genome Atlas ◽  
Tumor Type ◽  
Metastatic Progression ◽  
Statistical Machine Learning ◽  
Primary Tumors ◽  
Site Specific ◽  
Tissue Specific

Abstract Background & Aims Cancer metastasis into distant organs is an evolutionarily selective process. A better understanding of the driving forces endowing proliferative plasticity of tumor seeds in distant soils is required to develop and adapt better treatment systems for this lethal stage of the disease. To this end, we aimed to utilize transcript expression profiling features to predict the site-specific metastases of primary tumors and second, to identify the determinants of tissue specific progression. Methods We used statistical machine learning for transcript feature selection to optimize classification and built tree-based classifiers to predict tissue specific sites of metastatic progression. Results We developed a novel machine learning architecture that analyzes 33 types of RNA transcriptome profiles from The Cancer Genome Atlas (TCGA) database. Our classifier identifies the tumor type, derives synthetic instances of primary tumors metastasizing to distant organs and classifies the site-specific metastases in 16 types of cancers metastasizing to 12 locations. Conclusions We have demonstrated that site specific metastatic progression is predictable using transcriptomic profiling data from primary tumors and that the overrepresented biological processes in tumors metastasizing to congruent distant loci are highly overlapping. These results indicate site-specific progression was organotropic and core features of biological signaling pathways are identifiable that may describe proliferative plasticity in distant soils.

scholarly journals Glycosylation of CD44 is implicated in CD44-mediated cell adhesion to hyaluronan.

1996 ◽  
Vol 132 (6) ◽  
pp. 1199-1208 ◽  
Author(s):  
A Bartolazzi ◽  
A Nocks ◽  
A Aruffo ◽  
F Spring ◽  
I Stamenkovic
Keyword(s):  
Cell Adhesion ◽  
Regulatory Mechanism ◽  
Cell Attachment ◽  
Glycosylation Site ◽  
Side Chain ◽  
Site Specific ◽  
Coated Substrate ◽  
Glycosylation Sites ◽  
Coated Surfaces

CD44-mediated cell adhesion to hyaluronate is controlled by mechanisms which are poorly understood. In the present work we examine the role of N-linked glycosylation and Ser-Gly motifs in regulating CD44-hyaluronate interaction. Our results show that treatment of a panel of human cell lines which constitutively express CD44 with the inhibitor of N-linked glycosylation tunicamycin results in the loss of attachment of these cells to hyaluronate-coated substrate. In contrast, treatment of the same cells with deoxymannojirimycin, which inhibits the conversion of high mannose oligosaccharides to complex N-linked carbohydrates, results in either no change or an increase in CD44-mediated adhesion to hyaluronate, suggesting that complex N-linked oligosaccharides may not be required for and may even inhibit CD44-HA interaction. Using human melanoma cells stably transfected with CD44 N-linked glycosylation site-specific mutants, we show that integrity of five potential N-linked glycosylation sites within the hyaluronate recognition domain of CD44 is critical for hyaluronate binding. Mutation of any one of these potential N-linked glycosylation sites abrogates CD44-mediated melanoma cell attachment to hyaluronate-coated surfaces, suggesting that all five sites are necessary to maintain the HA-recognition domain in the appropriate conformation. We also demonstrate that mutation of serine residues which constitute the four Ser-Gly motifs in the membrane proximal domain, and provide potential sites for glycosaminoglycan side chain attachment, impairs hyaluronate binding. Taken together, these observations indicate that changes in glycosylation of CD44 can have profound effects on its interaction with hyaluronic acid and suggest that glycosylation may provide an important regulatory mechanism of CD44 function.

Method for Characterizing Sulfated Glycoproteins in a Glycosylation Site-Specific Fashion, Using Ion Pairing and Tandem Mass Spectrometry

2006 ◽  
Vol 78 (4) ◽  
pp. 1181-1190 ◽  
Author(s):  
Janet Irungu ◽  
Dilusha S. Dalpathado ◽  
Eden P. Go ◽  
Hui Jiang ◽  
Hy-Vy Ha ◽  
...  
Keyword(s):  
Mass Spectrometry ◽  
Tandem Mass Spectrometry ◽  
Ion Pairing ◽  
Glycosylation Site ◽  
Tandem Mass ◽  
Site Specific

scholarly journals Glycosylation Site-Specific Analysis of Clade C HIV-1 Envelope Proteins

2009 ◽  
Vol 8 (9) ◽  
pp. 4231-4242 ◽  
Author(s):  
Eden P. Go ◽  
Qing Chang ◽  
Hua-Xin Liao ◽  
Laura L. Sutherland ◽  
S. Munir Alam ◽  
...  
Keyword(s):  
Envelope Proteins ◽  
Glycosylation Site ◽  
Site Specific ◽  
Specific Analysis ◽  
Clade C ◽  
Hiv 1

scholarly journals Floxed exon (Flexon): A flexibly positioned stop cassette for recombinase-mediated conditional gene expression

2022 ◽  
Vol 119 (3) ◽  
pp. e2117451119
Author(s):  
Justin M. Shaffer ◽  
Iva Greenwald
Keyword(s):  
Gene Expression ◽  
Fluorescent Protein ◽  
Specific Protein ◽  
Specific Expression ◽  
Control Of Gene Expression ◽  
Site Specific ◽  
Tissue Specific ◽  
Endogenous Gene ◽  
Conditional Gene Expression ◽  
Stop Cassette

Conditional gene expression is a powerful tool for genetic analysis of biological phenomena. In the widely used “lox-stop-lox” approach, insertion of a stop cassette consisting of a series of stop codons and polyadenylation signals flanked by lox sites into the 5′ untranslated region (UTR) of a gene prevents expression until the cassette is excised by tissue-specific expression of Cre recombinase. Although lox-stop-lox and similar approaches using other site-specific recombinases have been successfully used in many experimental systems, this design has certain limitations. Here, we describe the Floxed exon (Flexon) approach, which uses a stop cassette composed of an artificial exon flanked by artificial introns, designed to cause premature termination of translation and nonsense-mediated decay of the mRNA and allowing for flexible placement into a gene. We demonstrate its efficacy in Caenorhabditis elegans by showing that, when promoters that cause weak and/or transient cell-specific expression are used to drive Cre in combination with a gfp(flexon) transgene, strong and sustained expression of green fluorescent protein (GFP) is obtained in specific lineages. We also demonstrate its efficacy in an endogenous gene context: we inserted a flexon into the Argonaute gene rde-1 to abrogate RNA interference (RNAi), and restored RNAi tissue specifically by expression of Cre. Finally, we describe several potential additional applications of the Flexon approach, including more precise control of gene expression using intersectional methods, tissue-specific protein degradation, and generation of genetic mosaics. The Flexon approach should be feasible in any system where a site-specific recombination-based method may be applied.

scholarly journals Selective elongation of the oligosaccharide attached to the second potential glycosylation site of yeast exoglucanase: effects on the activity and properties of the enzyme

1994 ◽  
Vol 304 (3) ◽  
pp. 917-922 ◽  
Author(s):  
R D Basco ◽  
L M Hernández ◽  
M D Muñox ◽  
I Olivero ◽  
E Andaluz ◽  
...  
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Thermal Inactivation ◽  
Glycosylation Site ◽  
Translation Product ◽  
Site Specific ◽  
Potential Glycosylation Site ◽  
Glycosylation Sites ◽  
Conformational Constraints ◽  
Specific Elongation ◽  
Protein Portion

Three exoglucanases (Exgs), ExgIa, ExgIb and Exg325, are secreted by Saccharomyces cerevisiae cells. They share a common protein portion with two potential glycosylation sites (sequons) but differ in the amount of N-linked carbohydrate [Basco, R.D., Muñoz, M.D., Hernández, L.M., Váquez de Aldana, C. and Larriba, G. (1993) Yeast 9, 221-234]. ExgIb contains two short oligosaccharides attached to asparagines (Asn) 165 and 325 of the primary translation product [Hernández, L.M., Olivero, I., Alvarado, E. and Larriba, G. (1992) Biochemistry 31, 9823-9831]. Exg325 carries a single, short oligosaccharide bound to Asn325 whereas ExgIa has at least one large oligosaccharide, since it has not been produced by mutant mnn9. To address the question of the origin of ExgIa, both sequons were individually mutated by substituting Gln for Asn. An ExgIa-like isoenzyme was still secreted by mutant Exg165 but not by mutant Exg325. Additional studies on sequential deglycosylation of ExgIa with endo-beta-N-acetylglucosaminidase H (endo H), the susceptibility of both oligosaccharides to the endoglycosidase, and analysis of the presence of GlcNAc at both asparagine residues after total deglycosylation with endo H, indicated that ExgIa contained two oligosaccharides, a short one bound to Asn165 and a large one bound to Asn325, and, accordingly, originated from ExgIb. The elongation of the second oligosaccharide did not result in a higher stability towards thermal inactivation or unfolding, or in an increased resistance to proteases as compared with ExgIb; however, the affinity of the enzyme towards laminarin decreased by 50%. This site-specific elongation occurred in the oligosaccharide that was less susceptible to endo H, indicating that these properties are determined by different conformational constraints.

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