POPS (phosphatidylglycerolphosphate synthase) is the most highly regulated enzyme in the yeast CL (cardiolipin) biosynthetic pathway. Therefore, we focused on this enzyme to characterize CL regulation and function. We solubilized POPS from the mitochondrial membrane of S. pombe using chaotropic agent and Triton X-100. The solubilized enzyme was further purified using gel filtration, ionic exchange chromatography, salt-induced phase separation, hydroxylapatite chromatography and affinity chromatography. The procedure yielded a homogeneous protein preparation; evidenced by both SDS-PAGE and agarose IEF under nondenaturing conditions. The identification of the enzyme was also proved by the fact that POPS activity could be steadily recovered from the agarose IEF gel slice containing the protein spot. Cloning of the PGPS gene by reverse genetics is in progress.
|Original language||English (US)|
|State||Published - Dec 1 1996|
ASJC Scopus subject areas
- Agricultural and Biological Sciences (miscellaneous)
- Biochemistry, Genetics and Molecular Biology(all)
- Cell Biology