Linked Life Data

2165501

Source:http://linkedlifedata.com/resource/pubmed/id/2165501

Statements in which the resource exists as a subject.
PredicateObject
rdf:type
lifeskim:mentions
pubmed:issue
22
pubmed:dateCreated
1990-9-4
pubmed:abstractText
The tricarboxylate transporter has been purified in reconstitutively active form from rat liver mitochondria. The transporter was extracted from mitoplasts with Triton X-114 in the presence of cardiolipin and citrate and was then purified by sequential chromatography on hydroxylapatite, Matrex Gel Orange A, Matrex Gel Blue B, and Affi-Gel 501. Analysis of the purified material via sodium dodecyl sulfate-polyacrylamide gel electrophoresis indicated the presence of one main protein band with an apparent molecular mass of 32.5 kDa. Upon incorporation into phospholipid vesicles, the purified transporter catalyzed a 1,2,3-benzenetricarboxylate-sensitive citrate/citrate exchange with a specific transport activity of 3240 nmol/4 min/mg of protein. This value was enhanced 831-fold with respect to the starting material. Substrate competition studies indicated that the reconstituted transport could be substantially inhibited by isocitrate, malate, and phosphoenolpyruvate, but not by alpha-ketoglutarate, succinate, malonate, pyruvate, or inorganic phosphate. Moreover, in addition to 1,2,3-benzenetricarboxylate, the reconstituted exchange was sensitive to the anion transport inhibitor n-butylmalonate but was insensitive to phenylsuccinate, alpha-cyano-4-hydroxycinnamate, and carboxyatractyloside. Finally, studies with covalent modifying agents indicated the purified transporter was inhibited by sulfhydryl reagents and by diethyl pyrocarbonate, 2,3-butanedione, phenylglyoxal, and pyridoxal 5-phosphate. In conclusion, these studies describe the first procedure to yield a highly purified tricarboxylate transport protein that both displays a high specific transport activity and can be obtained in quantities that readily enable further structural as well as functional studies. Based on its substrate specificity and inhibitor sensitivity, the purified 32.5-kDa protein appears to represent the complete tricarboxylate transport system found in rat liver mitochondria. Finally, new information is presented concerning the effect of covalent modifying reagents on the function of this transporter.
pubmed:grant
pubmed:language
eng
pubmed:journal
pubmed:citationSubset
IM
pubmed:chemical
pubmed:status
MEDLINE
pubmed:month
Aug
pubmed:issn
0021-9258
pubmed:author
pubmed:issnType
Print
pubmed:day
5
pubmed:volume
265
pubmed:owner
NLM
pubmed:authorsComplete
Y
pubmed:pagination
13379-85
pubmed:dateRevised
2007-11-14
pubmed:meshHeading
pubmed:year
1990
pubmed:articleTitle
Purification and characterization of the reconstitutively active tricarboxylate transporter from rat liver mitochondria.
pubmed:affiliation
Department of Pharmacology, College of Medicine, University of South Alabama, Mobile 36688.
pubmed:publicationType
Journal Article, Research Support, U.S. Gov't, P.H.S.