BiP and PDI cooperate in the oxidative folding of antibodies in vitro

Immunoglobulin heavy chain binding protein (BiP), a member of the Hsp70 cha perone family, and the oxidoreductase protein-disulfide isomerase (PDI) pla y an important role in the folding and oxidation of proteins in the endopla smic reticulum. However, it was not clear whether both cooperate in this pr ocess. We show here that BiP and PDI act synergistically in the in vitro fo lding of the denatured and reduced Fab fragment. Several ATP-dependent cycl es of binding, release, and rebinding of the unfolded antibody chains by Ri p are required for efficient reactivation. Our data suggest that in the abs ence of BiP unfolded antibody chains collapse rapidly upon refolding, rende ring cysteine side chains inaccessible for PDI. BiP binds the unfolded poly peptide chains and keeps them in a conformation in which the cysteine resid ues are accessible for PDI. These findings support the idea of a network of folding helper proteins in the endoplasmic reticulum, which makes this org anelle a dedicated protein-processing compartment.