Peroxiredoxin 6 homodimerization and heterodimerization with glutathione S-transferase pi are required for its peroxidase but not phospholipase A2 activity.

Peroxiredoxin 6 (Prdx6) is a unique 1-Cys member of the peroxiredoxin family with both GSH peroxidase and phospholipase A2 (PLA2) activities. It is highly expressed in the lung where it plays an important role in antioxidant defense and lung surfactant metabolism. Glutathionylation of Prdx6 mediated by its heterodimerization with GSH S-transferase π (πGST) is required for its peroxidatic catalytic cycle.

Ubiquitin is a novel substrate for human insulin-degrading enzyme.

Insulin-degrading enzyme (IDE) can degrade insulin and amyloid-β, peptides involved in diabetes and Alzheimer's disease, respectively. IDE selects its substrates based on size, charge, and flexibility. From these criteria, we predict that IDE can cleave and inactivate ubiquitin (Ub).

Insulin-degrading enzyme modulates the natriuretic peptide-mediated signaling response.

Natriuretic peptides (NPs) are cyclic vasoactive peptide hormones with high therapeutic potential. Three distinct NPs (ANP, BNP, and CNP) can selectively activate natriuretic peptide receptors, NPR-A and NPR-B, raising the cyclic GMP (cGMP) levels. Insulin-degrading enzyme (IDE) was found to rapidly cleave ANP, but the functional consequences of such cleavages in the cellular environment and the molecular mechanism of recognition and cleavage remain unknown.

Protective role of Cys-178 against the inactivation and oligomerization of human insulin-degrading enzyme by oxidation and nitrosylation.

Insulin-degrading enzyme (IDE), a 110-kDa metalloendopeptidase, hydrolyzes several physiologically relevant peptides, including insulin and amyloid-beta (Abeta). Human IDE has 13 cysteines and is inhibited by hydrogen peroxide and S-nitrosoglutathione (GSNO), donors of reactive oxygen and nitrogen species, respectively. Here, we report that the oxidative burst of BV-2 microglial cells leads to oxidation or nitrosylation of secreted IDE, leading to the reduced activity.

Structure, function, and regulation of insulin-degrading enzyme.

The short half-life of insulin in the human body (4-6 min) prompted the search and discovery of insulin-degrading enzyme (IDE), a 110-kDa metalloprotease that can rapidly degrade insulin into inactive fragments. Genetic and biochemical evidence accumulated in the last sixty years has implicated IDE as an important physiological contributor in the maintenance of insulin levels. Recent structural and biochemical analyses reveal the molecular basis of how IDE uses size and charge distribution of the catalytic chamber and structural flexibility of substrates to selectively recognize and degrade insulin, as well as the regulatory mechanisms of this enzyme.

Molecular bases for the recognition of short peptide substrates and cysteine-directed modifications of human insulin-degrading enzyme.

Insulin degrading enzyme (IDE) utilizes a large catalytic chamber to selectively bind and degrade peptide substrates such as insulin and amyloid beta (Abeta). Tight interactions with substrates occur at an exosite located approximately 30 A away from the catalytic center that anchors the N-terminus of substrates to facilitate binding and subsequent cleavages at the catalytic site. However, IDE also degrades peptide substrates that are too short to occupy both the catalytic site and the exosite simultaneously.

Characterization of the complex of glutathione S-transferase pi and 1-cysteine peroxiredoxin.

Glutathione S-transferase pi has been shown to reactivate 1-cysteine peroxiredoxin (1-Cys Prx) by formation of a complex [L.A. Ralat, Y.

Identification of tyrosine 79 in the tocopherol binding site of glutathione S-transferase pi.

Alpha-tocopherol, the most abundant form of vitamin E present in humans, is a noncompetitive inhibitor of glutathione S-transferase pi (GST pi), but its binding site had not been located. Tocopherol iodoacetate (TIA), a reactive analogue, produces a time-dependent inactivation of GST pi to a limit of 25% residual activity. The rate constant for inactivation, k(obs), exhibits a nonlinear dependence on reagent concentration, with K(I) = 19 microM and k(max) = 0.

Direct evidence for the formation of a complex between 1-cysteine peroxiredoxin and glutathione S-transferase pi with activity changes in both enzymes.

Glutathione S-transferase pi (GST pi) has been shown to reactivate oxidized 1-cysteine peroxiredoxin (1-Cys Prx, Prx VI, Prdx6, and AOP2). We now demonstrate that a heterodimer complex is formed between 1-Cys Prx with a C-terminal His6 tag and GST pi upon incubation of the two proteins at pH 8.0 in buffer containing 20% 1,6-hexanediol to dissociate the homodimers, followed by dialysis against buffer containing 2.

Glutathione S-transferase Pi has at least three distinguishable xenobiotic substrate sites close to its glutathione-binding site.

Benzyl isothiocyanate (BITC), present in cruciferous vegetables, is an efficient substrate of human glutathione S-transferase P1-1 (hGST P1-1). BITC also acts as an affinity label of hGST P1-1 in the absence of glutathione, yielding an enzyme inactive toward BITC as substrate. As monitored by using BITC as substrate, the dependence of k of inactivation (K(I)) of hGST P1-1 on [BITC] is hyperbolic, with K(I) = 66 +/- 7 microM.

Monobromobimane occupies a distinct xenobiotic substrate site in glutathione S-transferase pi.

Monobromobimane (mBBr), functions as a substrate of porcine glutathione S-transferase pi (GST pi): The enzyme catalyzes the reaction of mBBr with glutathione. S-(Hydroxyethyl)bimane, a nonreactive analog of monobromobimane, acts as a competitive inhibitor with respect to mBBr as substrate but does not affect the reaction of GST pi with another substrate, 1-chloro-2,4-dinitrobenzene (CDNB). In the absence of glutathione, monobromobimane inactivates GST pi at pH 7.