Purification and CharacteriZation of Golgi Membrane- Bound Nucleoside Diphosphatase from Suspension - Cultured CellS Of Sycamore (Acerpseudoplatanus L.)
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چکیده
Inosine diphosphatase (IDPase) isoforms associated with Golgi membranes were studied in sycamore cell culture. These enzyme isoforms were solubilized with Triton X-100 and purified by chromatography using DEAE-Toyopearl and SOURCES columns. The isoforms were separated into two distinguishable fractions (peak I and 2) by SOURCE S column c.hromatography. Furthermore the peak l contained at least two isoform bands detected by nativePAGE analysis The apparent molecular sizes of these three isoforms were estimated by both gel filtration and SDSPAGE to be 50 kDa, indicating that the Golgi membrane bound IDPase has a monomeric structure. These IDPase isoforms required divalent cations (Ca~7+. Mg2+, C02+, Mn~~+) for their hydrolyzing activity, and were inhibited by ATP. IDP, UDP, and GDP were effective substrates for these enzymes. It is clearly indicated that the sycamore Golgi membrane bound IDPase is a nucleoside diphosphatase.
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