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|Title:||Purification and characterization of a type II phosphatidylinositol 4-kinase from rat spleen and comparison with a Ptdlns 4-kinase from lymphocytes|
Fernandis, Aaron Z
|Abstract:||A Ptdlns 4-kinase from rat spleen particulate fraction was purified to homogeneity and its molecular properties were compared with a Ptdlns 4-kinase from splenic lymphocytes. The enzyme activity was solubilized from spleen particulate fraction with Triton X-100 and chromatographed sequentially on phosphocellulose, DEAE-sephacel, heparin acrylamide and hydroxyapatite columns. The purified enzyme preparation showed a 55 kDa band on SDS-PAGE with silver staining. Renaturation of the enzyme activity from SDS-PAGE showed that it comigrated with the 55 kDa protein. Characterization of the enzyme showed that it was a type II Ptdlns 4-kinase. Polyclonal antibodies raised against Ptdlns 4-kinase inhibited the enzyme activity in in vitro assays. Analysis of adult rat tissue particulate fractions on immunoblots showed restricted immunoreactivity among Ptdlns 4-kinases. However, the immunoreactivity is conserved in lymphoid tIssues from mouse to human, suggesting that lymphoid tissue has a distinct Ptdlns 4-kinase. Activation of rat splenocytes with Con A showed two fold in crease in Ptdlns 4-kinase activity. Comparison of Ptdlns 4-kinases from spleen and splenic lymphocytes showed identical chromatographic behaviour, molecular mass, immunoreactivity, Km values for Ptdlns and inhibition by adenosine.|
|ISSN:||0975-0959 (Online); 0301-1208 (Print)|
|Appears in Collections:||IJBB Vol.36(1) [February 1999]|
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