Linked Life Data

8132660

Source:http://linkedlifedata.com/resource/pubmed/id/8132660

Statements in which the resource exists as a subject.
PredicateObject
rdf:type
lifeskim:mentions
pubmed:issue
12
pubmed:dateCreated
1994-4-21
pubmed:abstractText
A novel form of an ATP-regulated, oligomeric, Ca(2+)-independent phospholipase A2 (iPLA2) has been purified from the cytosol of the murine macrophage-like cell line P388D1. The purification procedure included ammonium sulfate precipitation and sequential column chromatography on octyl-Sepharose, ATP-agarose, Mono Q fast protein liquid chromatography (FPLC), and hydroxyapatite FPLC. The resulting enzyme preparation was purified over 400,000-fold with a final specific activity of approximately 5 mumol/min/mg using a mixed micelle assay system of Triton X-100 and dipalmitoyl phosphatidylcholine (PC). The purified enzyme was Ca(2+)-independent and did not show a preference for either sn-2 arachidonic acid or sn-1 alkyl-ether containing phospholipids when utilizing mixed micelles as substrate. It was found to hydrolyze dipalmitoyl-PC approximately 4-fold faster than 1-palmitoyl-2-arachidonyl-PC and approximately 15-fold faster than 1-O-hexadecyl-2-arachidonyl-PC. Triton X-100 increased the P388D1 iPLA2 activity with optimal activity found at a Triton/phospholipid molar ratio of 4:1. The purified enzyme was activated 2-6-fold by ATP as well as other di- and triphosphate nucleosides. This activation was sensitive to the concentration of Triton X-100 present in the assay. SDS-polyacrylamide gel electrophoresis carried out on the purified enzyme yielded a single major band at a molecular weight of about 80,000. However, radiation inactivation experiments, carried out on the cell homogenate, demonstrated a target size of 337 +/- 25 kDa, indicating that the catalytically active iPLA2 exists as a large oligomeric complex, either through self-aggregation or association of the enzyme with other proteins.
pubmed:grant
pubmed:language
eng
pubmed:journal
pubmed:citationSubset
IM
pubmed:chemical
pubmed:status
MEDLINE
pubmed:month
Mar
pubmed:issn
0021-9258
pubmed:author
pubmed:issnType
Print
pubmed:day
25
pubmed:volume
269
pubmed:owner
NLM
pubmed:authorsComplete
Y
pubmed:pagination
9227-33
pubmed:dateRevised
2007-11-15
pubmed:meshHeading
pubmed:year
1994
pubmed:articleTitle
Ca(2+)-independent cytosolic phospholipase A2 from macrophage-like P388D1 cells. Isolation and characterization.
pubmed:affiliation
Department of Chemistry, University of California, San Diego, La Jolla 92093-0601.
pubmed:publicationType
Journal Article, Research Support, U.S. Gov't, P.H.S., Research Support, Non-U.S. Gov't