Oxidative Folding - in Eukaryotes

In Eukaryotes

A very similar pathway is followed in eukaryotes, in which the protein relay consists of proteins with very analogous properties as those of the protein relay in Gram-negative bacteria. However, a major difference between prokaryotes and eukaryotes is found in the fact that the process of oxidative protein folding occurs in the endoplasmatic reticulum (ER) in eukaryotes. A second difference is that in eukaryotes, the use of molecular oxygen as a terminal electron acceptor is not linked to the process of oxidative folding through the respiratory chain as is the case in bacteria. In fact, one of the proteins involved in the oxidative folding process uses a flavin-dependent reaction to pass electrons directly to molecular oxygen.

A homolog of DsbA, called protein disulfide isomerase (PDI), is responsible for the formation of the disulfide bonds in unfolded eukaryotic proteins. This protein has two thioredoxine-like active sites, which both contain two cysteine residues. By transferring the disulfide bond between these two cysteine residues onto the folding protein it is responsible for the latter’s oxidation. In contrast to bacteria, where the oxidative and isomerization pathways are carried out by different proteins, PDI is also responsible for the reduction and isomerization of the disulfide bonds. In order for PDI to catalyse the formation of disulfide bonds in unfolded proteins, it has to be reoxidized. This is carried out by an ER membrane-associated protein, Ero1p, which is no homolog of DsbB. This Ero1p protein forms a mixed disulfide with PDI, which is resolved by a nucleophilic attack of the second cystein residue in one of the active sites of PDI. As result, oxidized PDI is obtained. Ero1p itself is oxidized by transferring electrons to molecular oxygen. As it is an FAD-binding protein, this transfer of electrons is strongly favoured when Ero1p is bound to FAD. Also a transport system that imports FAD into the ER lumen has been described in eukaryotes. Furthermore, it has been shown that the ability to reduce or rearrange incorrect disulfide bonds in missfolded proteins is provided by the oxidation of reduced glutathione (GSH) to oxidized glutathione (GSSG).