Squalene Synthetase

Studies on the properties of the reaction and the mechanism of conversion of farnesyl pyrophosphate to squalene have been carried out with purified yeast squalene synthetase. The general properties of the yeast enzyme are similar to those of the enzyme obtained from mammalian liver. A divalent metal ion (Mg++ or Mn++) is essential for the formation of presqualene pyrophosphate, but not for the conversion of this compound to squalene. However, the rate of the latter reaction is increased in the presence of Mg++. Yeast squalene synthetase activity is inhibited by concentrations above 50 mm of farnesyl pyrophosphate, by N-ethylmaleimide, and to a lesser extent by iodoacetamide. Hence it is evident that a sulfhydryl group or other nucleophile is required for enzyme activity. Yeast squalene synthetase has a maximum enzyme activity at pH 7.3 to 7.5. However, great differences in enzyme activity were found, depending on the buffer used. These differences are due in part to ionic strength, and possibly to a direct effect of phosphate ion on enzymatic activity. Initial velocity studies for the formation of squalene and presqualene pyrophosphate from farnesyl pyrophosphate have shown that the mechanism of the condensing reaction is ping-pong. Initial velocity kinetics and product inhibition studies for the reduction of presqualene pyrophosphate to squalene have shown that this reaction is sequential ordered. In the reduction reaction NADPH is the first substrate to bind to the enzyme, followed by presqualene pyrophosphate. The first product to be released is pyrophosphate, followed by squalene. The last product to leave the enzyme is NADP. A chemical mechanism for squalene synthesis is presented.