SAC1P MEDIATES THE ADENOSINE-TRIPHOSPHATE TRANSPORT INTO YEAST ENDOPLASMIC-RETICULUM THAT IS REQUIRED FOR PROTEIN TRANSLOCATION

Protein translocation into the yeast endo plasmic reticulum requires t he transport of ATP into the lumen of this organelle. Microsomal ATP t ransport activity was reconstituted into proteoliposomes to characteri ze and identify the transporter protein. A polypeptide was purified wh ose partial amino acid sequence demonstrated its identity to the produ ct of the SAC1 gene. Accordingly, microsomal membranes isolated from s trains harboring a deletion in the SAC1 gene (sac1 Delta) were found t o be deficient in ATP-transporting activity as well as severely compro mised in their ability to translocate nascent prepro-alpha-factor and preprocarboxypeptidase Y. Proteins isolated from the microsomal membra nes of a sac1 Delta strain were incapable of stimulating ATP transport when reconstituted into the in vitro assay system. When immunopurifie d to homogeneity and incorporated into artificial lipid vesicles. Sac1 p was shown to reconstitute ATP transport activity. Consistent with th e requirement for ATP in the lumen of the ER to achieve the correct fo lding of secretory proteins. the sac1 Delta strain was shown to have a severe defect in transport of procarboxypeptidase Y out of the ER and into the Golgi complex in vivo. The collective data indicate an intim ate role for Sac1p in the transport of ATP into the ER lumen.