INTRACELLULAR-DISTRIBUTION AND SOLUBILIZATION OF YEAST DOLICHYLPYROPHOSPHATASE

Yeast dolichylpyrophosphatase is associated with the membrane apparatus of yeast cells. The enzyme is localized primarily in the microsome fraction and in vacuoles; moreover, high specific activity of dolichylpyrophosphatase has been noted in the "light membrane" fraction, flotated in a ficoll gradient at high rates of sedimentation. Dolichylpyrophosphatase has been solubilized from the total yeast membrane fraction using the nonionic detergent Triion X-100 in the presence of 6 mM MgCl2 and 18 mM NaCl. The specific activity of the solubilized enzyme is more than 40 times as high as the activity in the initial protoplast lysate. EDTA substantially activates dolichylpyrophosphatase (by a factor of three), whereas orthophosphate and most divalent metal cations are strong inhibitors. The solubilized enzyme can be stored without any substantial loss of activity for six months at -10-degrees-C.