BiP and PDI cooperate in the oxidative folding of antibodies in vitro.
Immunoglobulin heavy chain binding protein (BiP), a member of the Hsp70 chaperone family, and the oxidoreductase protein-disulfide isomerase (PDI) play an important role in the folding and oxidation of proteins in the endoplasmic reticulum. However, it was not clear whether both cooperate in this process. We show here that BiP and PDI act synergistically in the in vitro folding of the denatured and reduced Fab fragment. Several ATP-dependent cycles of binding, release, and rebinding of the unfolded antibody chains by BiP are required for efficient reactivation. Our data suggest that in the absence of BiP unfolded antibody chains collapse rapidly upon refolding, rendering cysteine side chains inaccessible for PDI. BiP binds the unfolded polypeptide chains and keeps them in a conformation in which the cysteine residues are accessible for PDI. These findings support the idea of a network of folding helper proteins in the endoplasmic reticulum, which makes this organelle a dedicated protein-processing compartment.
Mayer
M
M
Institut für Organische Chemie & Biochemie, Technische Universität München, 85747 Garching, Germany.
Kies
U
U
Kammermeier
R
R
Buchner
J
J
D002352:Carrier Proteins / Q000737:chemistry / Q000378:metabolism*; D003545:Cysteine; D000091342:Endoplasmic Reticulum Chaperone BiP; D004926:Escherichia coli; D006360:Heat-Shock Proteins / Q000378:metabolism; D006801:Humans; D007128:Immunoglobulin Fragments / Q000737:chemistry / Q000378:metabolism*; D007143:Immunoglobulin Heavy Chains / Q000378:metabolism; D018832:Molecular Chaperones / Q000737:chemistry / Q000378:metabolism*; D010084:Oxidation-Reduction; D011485:Protein Binding; D019704:Protein Disulfide-Isomerases / Q000737:chemistry / Q000378:metabolism*; D017510:Protein Folding
D002352:Carrier Proteins; D000091342:Endoplasmic Reticulum Chaperone BiP; D006360:Heat-Shock Proteins; D007128:Immunoglobulin Fragments; D007143:Immunoglobulin Heavy Chains; D018832:Molecular Chaperones; D019704:Protein Disulfide-Isomerases; D003545:Cysteine