Linked Life Data

10361002

Source:http://linkedlifedata.com/resource/pubmed/id/10361002

Statements in which the resource exists as a subject.
PredicateObject
rdf:type
lifeskim:mentions
pubmed:issue
1
pubmed:dateCreated
1999-8-13
pubmed:abstractText
We have developed a nonradioactive method to assay UDP-Gal:beta-d-GlcNAcbeta1,4-galactosyltransferase (beta4GalT-I) enzymatic activity. Capillary electrophoresis combined with laser-induced fluorescence detection (CE-LIF) was employed to provide a baseline separation of FITC-conjugated O-GlcNAc-containing substrate peptides and galactose-capped product peptides, while at the same time allowing a level of detection in the low attomole range (10(-18)). The addition of 2 mM hexamethylene diamine to the borate-based capillary electrophoretic buffer modulated the electroosmotic flow, resulting in optimum separation of the glycopeptide product from reactant. beta4GalT-I activity was dependent upon the addition of both manganese and UDP-galactose. Using this assay, we show that two beta4GalT-I constructs, predicted to localize to different intracellular compartments, are enzymatically active when expressed in vitro using a rabbit reticulocyte transcription-translation system. The high sensitivity of product detection by CE-LIF in combination with in vitro transcription-translation is applicable to the facile determination of the enzymatic activity of other newly cloned glycosyltransferases.
pubmed:grant
pubmed:language
eng
pubmed:journal
pubmed:citationSubset
IM
pubmed:chemical
pubmed:status
MEDLINE
pubmed:month
Jun
pubmed:issn
0003-2697
pubmed:author
pubmed:copyrightInfo
Copyright 1999 Academic Press.
pubmed:issnType
Print
pubmed:day
15
pubmed:volume
271
pubmed:owner
NLM
pubmed:authorsComplete
Y
pubmed:pagination
36-42
pubmed:dateRevised
2007-11-14
pubmed:meshHeading
pubmed:year
1999
pubmed:articleTitle
Determination of beta1,4-galactosyltransferase enzymatic activity by capillary electrophoresis and laser-induced fluorescence detection.
pubmed:affiliation
School of Medicine, The Johns Hopkins University School of Medicine, Baltimore, Maryland 21207, USA.
pubmed:publicationType
Journal Article, In Vitro, Research Support, U.S. Gov't, P.H.S.