Evidence for the involvement of a 66 kDa membrane protein in the synthesis of sterolglucoside in Saccharomyces cerevisiae.

Abstract

The membrane-bound sterolglucoside synthase from the yeast Saccharomyces cerevisiae has been solubilized by nonionic detergent, Nonidet P-40, Triton X-100, and partially purified by DEAE-cellulose column chromatography and ammonium sulfate fractionation. SDS/PAGE of the purified fraction revealed the presence of two protein bands of molecular mass 66 kDa and 54 kDa. In an attempt to identify further the polypeptide chain of sterolglucoside synthase, the partially purified enzyme was treated with [di-125I]-5-[3-(p-azidosalicylamide)]allyl-UDPglucose, a photoactive analogue of UDP glucose, which is a substrate for this enzyme. Upon photolysis the 125I-labeled probe was shown to link covalently to the 66 kDa protein. The photoinsertion was competed out by the presence of unlabeled UDPglucose thus suggesting that this protein contains substrate binding site for UDPglucose. Since photoinsertion of the probe to protein of 66 kDa correlates with the molecular mass of the protein visualized upon enzyme purification we postulate that the 66 kDa protein is involved in sterolglucoside synthesis in yeast.

Properties of recombinant trehalose synthase from Deinococcus radiodurans expressed in Escherichia coli.

“…One of the reasons for trehalose applications is participation in stabilization of proteins and/or biological membranes. Probably trehalose can form amorphous glass which entraps biological molecules without essential changes to their native structure during desiccation, freezing, heating or oxidation (Crove & Crove, 2000;Roser, 1991;Benaroudi, 2001;Lenart et al, 1995). According to an alternate hypothesis trehalose can substitute water around biomolecules that provide hydrogen bonding network maintaining the structure of the protein.…”