Protein disulfides and protein disulfide oxidoreductases in hyperthermophiles.

Disulfide bonds are required for the stability and function of a large number of proteins. Recently, the results from genome analysis have suggested an important role for disulfide bonds concerning the structural stabilization of intracellular proteins from hyperthermophilic Archaea and Bacteria, contrary to the conventional view that structural disulfide bonds are rare in proteins from Archaea. A specific protein, known as protein disulfide oxidoreductase (PDO) is recognized as a potential key player in intracellular disulfide-shuffling in hyperthermophiles. The structure of this protein shows a combination of two thioredoxin-related units with low sequence identity which together, in tandem-like manner, form a closed protein domain. Each of these units contains a distinct CXXC active site motif. Due to their estimated conformational energies, both sites are likely to have different redox properties. The observed structural and functional characteristics suggest a relation to eukaryotic protein disulfide isomerase. Functional studies have revealed that both the archaeal and bacterial forms of this protein show oxidative and reductive activity and are able to isomerize protein disulfides. The physiological substrates and reduction systems, however, are to date unknown. The variety of active site disulfides found in PDOs from hyperthermophiles is puzzling. Nevertheless, the catalytic function of any PDO is expected to be correlated with the redox properties of its active site disulfides CXXC and with the distinct nature of its redox environment. The residues around the two active sites form two grooves on the protein surface. In analogy to a similar groove in thioredoxin, both grooves are suggested to constitute the substrate binding sites of PDO. The direct neighbourhood of the grooves and the different redox properties of both sites may favour sequential reactions in protein disulfide shuffling, like reduction followed by oxidation. A model for peptide binding by PDO is proposed to be derived from the analysis of crystal packing contacts mimicking substrate binding interactions. It is assumed, that PDO enzymes in hyperthermophilic Archaea and Bacteria may be part of a complex system involved in the maintenance of protein disulfide bonds. The regulation of disulfide bond formation may be dependent on a distinct interplay of thermodynamic and kinetic effects, including functional asymmetry and substrate-mediated protection of the active sites, in analogy to the situation in protein disulfide isomerase. Numerous questions related to the function of PDO enzymes in hyperthermophiles remain unanswered to date, but can probably successfully be studied by a number of approaches, such as first-line genetic and in vivo studies.