Structural Plasticity of the Thioredoxin Recognition Site of Yeast Methionine S-Sulfoxide Reductase Mxr1

Abstract

The methionine S-sulfoxide reductase MsrA catalyzes the reduction of methionine sulfoxide, a ubiquitous reaction depending on the thioredoxin system. To investigate interactions between MsrA and thioredoxin (Trx), we determined the crystal structures of yeast MsrA/Mxr1 in their reduced, oxidized, and Trx2-complexed forms, at 2.03, 1.90, and 2.70 Å , respectively. Comparative structure analysis revealed significant conformational changes of the three loops, which form a plastic "cushion" to harbor the electron donor Trx2. The flexible C-terminal loop enabled Mxr1 to access the methionine sulfoxide on various protein substrates. Moreover, the plasticity of the Trx binding site on Mxr1 provides structural insights into the recognition of diverse substrates by a universal catalytic motif of Trx. Thioredoxins (Trxs)2 are ubiquitous small thiol-disulfide exchange proteins that are involved in many important cellular processes such as reduction of methionine sulfoxide, ribonucleotide, and peroxide (1). These proteins have a highly conserved active site of CXXC motif. During the reaction, the first Cys attacks the intramolecular disulfide bond in the substrate protein, accompanying w ith the formation of an intermolecular disulfide intermediate. This mixed disulfide is subsequently attacked by the second Cys, resulting in release of the reduced substrate protein and the oxidized Trx.

Dissecting Molecular Interactions Involved in Recognition of Target Disulfides by the Barley Thioredoxin System

“…The A106 NH hydrogen bond may be important for reduction of PAPS reductase and methionine sulfoxide reductase because homologues of A106 in Trx complexes with these target proteins participate in similar intermolecular backbone−backbone hydrogen bonds as observed in the HvTrxh2−BASI complex. 7,8,10 While the role of this highly conserved alanine residue was not previously investigated by mutagenesis, the invariant preceding glycine residue (G92 of EcTrx1) has previously been identified as being critical for interactions with EcNTR and bacteriophage T7 DNA polymerase. 37 The side chain of HvTrxh2 M88 is important for efficient target disulfide reduction as demonstrated by the same order of decreasing activity found for HvTrxh2 (wt > M88L > M88A > M88G) toward BASI, Gpx, and HvNTR2 (Table 3).…”

“…5,6,8,9 In the recently determined structure of yeast Trx2 in complex with methionine sulfoxide reductase, the similarity is striking as three fully analogous antiparallel backbone−backbone hydrogen bonds are found. 10 The interactions in the HvTrxh2−BASI complex therefore appear to be characteristic of disulfide target recognition by Trxs.…”

A universal entropy-driven mechanism for thioredoxin–target recognition

“…There has been a long-standing interest in identifying the primary forces involved in recognition of multiple target proteins by Trx. Several X-ray structures for covalent Trx-protein (or peptide) complexes have been reported (16)(17)(18). Although the static structures afforded by X-ray crystallography can provide details about enthalpy of interaction at atomic resolution, they seldom provide information regarding entropic forces, thus necessitating the use of alternative biophysical methods for complete thermodynamic characterization of protein-protein interactions (28).…”

“…However, the molecular mechanism underlying this process, particularly in the absence of a signature Trx-binding sequence or structural motif, has remained largely unknown. Periodic efforts have been made to study individual Trx-target interactions through X-ray crystallography (16)(17)(18). In these studies, the Trx-protein complex has been trapped via the mixed disulfide that forms between the target and a Trx mutant, in which the resolving cysteine is mutated to an alanine or serine.…”

Proteome and fermentative parameters of Saccharomyces cerevisiae CAT-1 under Very High Gravity Fermentation (VHGF) using sugarcane juice

“…This enzyme is related to both the peptidyl-prolyl isomerization during protein folding or conformational changes and the meiosis to promote an efficient sporulation when associated with other two proteins from the histone deacetylase complexes and in the glucose-stimulated transport of fructose-1,6-bisphosphatase into Vid vesicles [38,39]. The protein Mxr1-Trx2 is a methionine S-sulfoxide reductase that uses thioredoxin as electron donor [40,41]. Both thioredoxin-1 (Trx1) and thioredoxin-2 (Trx2) were previously identified in the quantitative proteome analysis of S. cerevisiae CAT-1, constituting important elements in the stress tolerance observed for this yeast [2].…”