Linked Life Data

11067750

Source:http://linkedlifedata.com/resource/pubmed/id/11067750

Statements in which the resource exists as a subject.
PredicateObject
rdf:type
lifeskim:mentions
pubmed:issue
2
pubmed:dateCreated
2001-3-8
pubmed:abstractText
A nonradioactive assay for the investigation of phospholipase D (PLD) activity in cardiac membranes has been developed. A fluorescent derivative of phosphatidylcholine [2-decanoyl-1-(O-(11-(4,4-difluoro-5,7-dimethyl-4-bora-3a, 4a-diaza-s-indacene-3proprionyl)amino) undecyl) sn-glycero-3-phosphocholine] was utilized as substrate in an in vitro PLD-catalyzed transphosphatidylation reaction utilizing ethanol as second substrate. Unreacted phosphatidylcholine and the products of phospholipase activity (PEtOH, phosphatidylethanol; PA, phosphatidic acid; DAG, diacylglycerol) were separated by a binary gradient HPLC system and detected by fluorometry. The detection limit of this assay is approximately 0.6 pmol PEtOH. The reaction proceeded at a linear rate for up to 45 min and increased linearly with increasing amounts of rat cardiac membrane protein in a range of 0.625 microg up to at least 25 microg. In the presence of potassium fluoride, formation of fluorescent PA increased at the expense of DAG generation, demonstrating the presence of PA phosphohydrolase activity in rat cardiac membranes. PEtOH formation was unchanged in the presence of the PA phosphohydrolase inhibitor, indicating that the phosphatidylalcohol is not subject to further metabolism by this enzyme. Activation of protein kinase C by phorbol ester significantly increased PLD activity in cardiac membranes. This assay proved to be sensitive for accurate and rapid assessment of PLD activity in cardiac membranes permitting further characterization of the regulation of PLD signal transduction in the heart.
pubmed:language
eng
pubmed:journal
pubmed:citationSubset
IM
pubmed:chemical
pubmed:status
MEDLINE
pubmed:month
Nov
pubmed:issn
0003-2697
pubmed:author
pubmed:copyrightInfo
Copyright 2000 Academic Press.
pubmed:issnType
Print
pubmed:day
15
pubmed:volume
286
pubmed:owner
NLM
pubmed:authorsComplete
Y
pubmed:pagination
277-81
pubmed:dateRevised
2006-11-15
pubmed:meshHeading
pubmed:year
2000
pubmed:articleTitle
A HPLC-fluorescence detection method for determination of cardiac phospholipase D activity in vitro.
pubmed:affiliation
Medizinische Klinik II, Medizinische Universität zu Lübeck, Lübeck, Germany.
pubmed:publicationType
Journal Article, In Vitro