The origami of thioredoxin-like folds

Abstract

Origami is the Japanese art of folding a piece of paper into complex shapes and forms. Much like origami of paper, Nature has used conserved protein folds to engineer proteins for a particular task. An example of a protein family, which has been used by Nature numerous times, is the thioredoxin superfamily. Proteins in the thioredoxin superfamily are all structured with a beta-sheet core surrounded with alpha-helices, and most contain a canonical CXXC motif. The remarkable feature of these proteins is that the link between them is the fold; however, their reactivity is different for each member due to small variations in this general fold as well as their active site. This review attempts to unravel the minute differences within this protein family, and it also demonstrates the ingenuity of Nature to use a conserved fold to generate a diverse collection of proteins to perform a number of different biochemical tasks.

Figure 1.

Japanese origami. Paper is carefully folded into complex angles using different parts of the paper incorporated with delicate creases. Manipulation of the paper results in a functional representation of an object of shape. In this case, paper was folded to create a replica of a dinosaur. Proteins are also folded with careful creases, creating a functionally active biomolecule.

Figure 2.

Structure of oxidized thioredoxin. The thioredoxin fold, named for the founding member of the family thioredoxin, is a protein fold consisting of four β-sheets surrounded by three α-helices. The thioredoxin family of protein is typically involved in redox active reactions and usually contains a C-X-X-C motif, where X is any amino acid.

Figure 3.

Ribbon structures of representative members for the thioredoxin family. Members of the thioredoxin family have a characteristic fold with β-sheets surrounded by α-helices.

Figure 4.

Typical architecture of thioredoxin-like proteins. The figure displays the two-dimensional representation of the thioredoxin fold in representative proteins in the thioredoxin super family. β-Sheets are drawn as arrows and α-helices are drawn in boxes. A typical thioredoxin fold is shown where two parallel β-sheets are intervened by an α-helix. In addition, between the N-terminal and C-terminal portion is an α-helix that connects these two domains. Helices that are not part of the thioredoxin fold are drawn colored in white.

Figure 5.

Mechanism of dithiol and monothiol reactivities of glutaredoxin. (A) (1) Reduced glutaredoxin first seeks a substrate that has a disulfide bond; (2) with a reduced cysteine, glutaredoxin attacks the disulfide in the target protein resulting in a mixed disulfide; (3–4) with a deprotonated C-terminal cysteine, glutaredoxin attacks the mixed disulfide, which results in a reduced substrate protein and a completely oxidized glutaredoxin; (5) Glutaredoxin is then reduced to return back into its active form. (B) (1) Glutaredoxin starts off as a reduced species and attacks a protein that is glutathionylated on a cysteine. (2) Glutaredoxin becomes glutathionylated and the substrate protein becomes reduced with a free cystine. (3) A reduced glutathione molecule then attacks the mixed disulfide in glutaredoxin. (4) Glutaredoxin returns back into its reduced conformation and oxidized glutathione disulfide is formed.

Figure 6.

Mechanism of disulfide formation and isomerization in the bacterial periplasm. (A) Disulfide bonds are generated by the Dsb family of proteins in the bacterial periplasm. First, DsbA, a thioredoxin-like protein, has a catalytic disulfide and seeks to donate its disulfide to newly translocated proteins, resulting in a reduced DsbA and oxidized substrate protein. For the system to be catalytic, a membrane protein DsbB oxidizes DsbA. DsbB gains its oxidizing equivalents from the electron transport chain with the final electron acceptor oxygen. (B) DsbA is a potent disulfide oxidase and has the potential to incorporate incorrect disulfides. Another pathway exists to correct incorrect disulfides formed by DsbA or another exogenous disulfide source. DsbC is a thioredoxin-like protein that has reduced cysteines. These reduced cysteines accept a disulfide from a substrate protein. Maintaining DsbC in the reduced form is accomplished by the membrane protein DsbD. DsbD gains its reducing equivalents from the cytosolic protein thioredoxin, which ultimately gains its reduced equivalents from NADPH.

Figure 7.

Generalized mechanisms of atypical 2-Cys, typical 2-Cys, and 1-Cys peroxiredoxins. (A) In the typical 2-Cys peroxiredoxins, a homodimer, where one monomer has the active peroxidatic site cysteine and the other having the resolving cysteine. The peroxidatic becomes oxidized to a sulfenic acid, thereby reducing hydrogen peroxide to water. The resolving cysteine now attacks the peroxidatic cysteine to form an intersubunit disulfide bond. The disulfide is reduced from cytosolic reductants such as thioredoxin. (B) In the atypical 2-Cys peroxiredoxins, the mechanism of action is very similar to the typical 2-Cys peroxiredoxins except that both the peroxidatic and the resolving cysteines are both on the same monomer, thereby eliminating the need for two protein monomers. (C) The 1-Cys peroxiredoxins function differently than the 2-Cys peroxiredoxins. These peroxiredoxins interact with the peroxide directly and form a sulfenic acid intermediate on the peroxidatic cysteine. The 2-Cys peroxiredoxins then interact with a thiol reducing source such as thioredoxin or glutaredoxin and become reduced, releasing water and regenerating the reduced 1-Cys peroxiredoxin. (Derived from Wood et al. 2003.)