Formation of an intrachain disulfide bond on nascent immunoglobulin light chains.

Immunoglohulin light chains are comprised of two compact globular domains, each of which contains approqiimately 110 amino acid residues and a single intrachain disulfide bond. Using ion-exchange chromatography to purify peptidyl-tRNA complexes, we have analyzed the in vivo formation of intrachain disulfide bonds on nascent MPC 11 light chain polypeptides. We have demonstrated the presence of the correct intrachain disulfide loop (Cys 35-Cys 100) on some nascent light chains of approximately 15,500 daltons in mass. Eighty-five per cent of nascent light chains in a 15,500to l&000-dalton fraction and all nascent light chains larger than 18,000 daltons have formed this first domain intrachain disulfide loop. These results suggest that this disulfide loop (Cys 35-Cys 100) is formed quantitatively in less than 1 s after Cys 100 passes through the membrane into the cisterna of the endoplasmic reticulum. The intrachain disulfide loop in the second or constant region domain of light chain (Cys 146-Cys 206) is not formed on nascent polypeptides because one of the cysteine residues (Cys 206) involved in this disulfide bond does not pass through the endoplasmic reticulum membrane prior to chain completion and release from the ribosome. No incorrect intrachain disulfide bonds are detected on nascent light chains. These results indicate that in vivo proteins initiate folding sequentially from the NHz-terminal end to the COOH-terminal end as growing nascent chains prior to release from the ribosome. In view of the fact that the NHP-terminal regions of a nascent polypeptide have a longer time to fold than the more COOH-terminal regions, it is likely that the folding of a denatured completed polypeptide in vitro is not generally an accurate model of in vivo protein folding.