Misfolded polypeptides within the lumen of the endoplasmic reticulum are recognized and retrograde translocated to the cytoplasm for degradation by proteasomes. This retrograde translocation event is dependent upon Sec61p and is reductant sensitive, implying the importance of a disulfide bonded species. The proposed research will elucidate the reductant sensitive disulfide bond, and investigate the possibility that ER luminal protein disulfide isomerases promote disulfide bond isomerizations of Sec61p to produce dislocation specific channels. In addition, intermediates of the dislocation reaction will be engineered by the construction of chimeric proteins designed to become arrested shortly after the initiation of dislocation. Physical properties and dimensions of the dislocation channel will be measured by fluorescence spectroscopy utilizing the developed intermediates of the dislocation reaction.
