Solubilization and Purification of tram-Farnesyl

A trans-farnesyl pyrophosphate-squalene synthetase has been isolated in a soluble form from yeast extracts and purified 45fold. The molecular weight of the enzyme estimated from sucrose density gradient centrifugation and gel filtration chromatography is 426,000. Solubilization of the squalene synthetase is achieved with deoxycholate. Treatment with the detergent markedly lowers squalene synthetase activity but when deoxychola$e is removed by Amberlite XAD-2, the soluble enzyme regains full activity. Such synthetase preparations are relatively labile. They can be stabilized by glycerol and Z-mercaptoethanol. Both TPNH and DPNH serve as electron donors for the squalene synthetase. Their K, values are 122 pM and 310 pM, respectively. The two pyridine nucleotides differ somewhat in their effects on the Hill coefficient for the bimolecular condensation of farnesyl pyrophosphate to squalene. With DPNH the Hill slope is 2.0 and with TPNH 1.4. The purified synthetase catalyzes not only the formation of squalene from farnesyl pyrophosphate but also accumulates presqualene pyrophosphate (in the absence of pyridine nucleotide) and converts biosynthetic presqualene pyrophosphate to squalene.