MEMBRANE GLYCOPROTEIN FOLDING, OLIGOMERIZATION AND INTRACELLULAR-TRANSPORT - EFFECTS OF DITHIOTHREITOL IN LIVING CELLS

Using influenza hemagglutinin (HA0) and vesicular stomatitis virus G p rotein as model proteins, we have analyzed the effects of dithiothreit ol (DTT) on conformational maturation and transport of glycoproteins i n the secretory pathway of living cells. While DTT caused reduction of folding intermediates and misfolded proteins in the endoplasmic retic ulum (ER), it did not affect molecules that had already acquired a mat ure trimeric conformation, whether present in the ER or elsewhere. The conversion to DTT resistance was therefore a pre-Golgi event. Reducti on of folding intermediates was dependent on the intactness of the ER and on metabolic energy, suggesting cooperativity between DTT and ER f olding factors. DTT did not inhibit most cellular functions, including ATP synthesis and protein transport within the secretory pathway. The results established DTT as an effective tool for analyzing the foldin g and compartmental distribution of proteins with disulfide bonds.