Linked Life Data

10910972

Source:http://linkedlifedata.com/resource/pubmed/id/10910972

Statements in which the resource exists as a subject.
PredicateObject
rdf:type
lifeskim:mentions
pubmed:issue
7
pubmed:dateCreated
2000-9-27
pubmed:abstractText
The production of mice with genetic alterations in glycosyltransferases has highlighted the need to isolate and study complex mixtures of the major classes of oligosaccharides (glycans) from intact tissues. We have found that nano-NMR spectroscopy of whole mixtures of N- and O-glycans can complement HPLC profiling methods for elucidating structural details. Working toward obtaining such glycan mixtures from mouse tissues, we decided to develop an approach to isolate not only N- and O-glycans, but also to separate out glycosphingolipids, glycosaminoglycans and glycosylphosphatidylinositol anchors. We describe here a comprehensive Glycan Isolation Protocol that is based primarily upon the physicochemical characteristics of the molecules, and requires only commonly available reagents and equipment. Using radiolabeled internal tracers, we show that recovery of each major class of glycans is as good or better than with conventional approaches for isolating individual classes, and that cross-contamination is minimal. The recovered glycans are of sufficient purity to provide a "glycoprofile" of a cell type or tissue. We applied this approach to compare the N- and O-glycans from wild type mouse tissues with those from mice genetically deficient in glycosyltransferases. N- and O-glycan mixtures from organs of mice deficient in ST6Gal-I (CMP-Sia:Galbeta1-4GlcNAc alpha2-6 sialyltransferase) were studied by the nano-NMR spectroscopy approach, showing no detectable alpha2-6-linked sialic acids. Thus, ST6Gal-I is likely responsible for generating most or all of these residues in normal mice. Similar studies indicate that this linkage is very rare in ganglioside glycans, even in wild-type tissues. In mice deficient in GalNAcT-8 (UDP-GalNAc:polypeptide O-Ser/Thr GalNAc transferase 8), HPLC profiling indicates that O-glycans persist in the thymus in large amounts, without a major change in overall profile, suggesting that other enzymes can synthesize the GalNAc-O-Ser/Thr linkage in this tissue. These results demonstrate the applicability of nano-NMR spectroscopy to complex glycan mixtures, as well as the versatility of the Glycan Isolation Protocol, which makes possible the concurrent examination of multiple glycan classes from intact vertebrate tissues.
pubmed:grant
pubmed:language
eng
pubmed:journal
pubmed:citationSubset
IM
pubmed:chemical
pubmed:status
MEDLINE
pubmed:month
Jul
pubmed:issn
0959-6658
pubmed:author
pubmed:issnType
Print
pubmed:volume
10
pubmed:owner
NLM
pubmed:authorsComplete
Y
pubmed:pagination
669-89
pubmed:dateRevised
2007-11-14
pubmed:meshHeading
pubmed:year
2000
pubmed:articleTitle
Exploring the glycan repertoire of genetically modified mice by isolation and profiling of the major glycan classes and nano-NMR analysis of glycan mixtures.
pubmed:affiliation
Departments of Medicine and Cellular and Molecular Medicine, Howard Hughes Medical Institute, University of California San Diego, La Jolla 92093-0687, USA.
pubmed:publicationType
Journal Article, Research Support, U.S. Gov't, P.H.S., Research Support, Non-U.S. Gov't