Redox-active chemical chaperones exhibiting promiscuous binding promote oxidative protein folding under condensed sub-millimolar conditions.

Proteins form native structures through folding processes, many of which proceed through intramolecular hydrophobic effect, hydrogen bond and disulfide-bond formation. , protein aggregation is prevented even in the highly condensed milieu of a cell through folding mediated by molecular chaperones and oxidative enzymes. Chemical approaches to date have not replicated such exquisite mediation.

Functional Interplay between P5 and PDI/ERp72 to Drive Protein Folding.

P5 is one of protein disulfide isomerase family proteins (PDIs) involved in endoplasmic reticulum (ER) protein quality control that assists oxidative folding, inhibits protein aggregation, and regulates the unfolded protein response. P5 reportedly interacts with other PDIs via intermolecular disulfide bonds in cultured cells, but it remains unclear whether complex formation between P5 and other PDIs is involved in regulating enzymatic and chaperone functions. Herein, we established the far-western blot method to detect non-covalent interactions between P5 and other PDIs and found that PDI and ERp72 are partner proteins of P5.

Zinc-Dependent Oligomerization of Trigger Factor Chaperone.

trigger factor (TF) is a zinc-dependent molecular chaperone whose folding-arrest activity is regulated by Zn. However, little is known about the mechanism of zinc-dependent regulation of the TF activity. Here we exploit in vitro biophysical experiments to investigate zinc-binding, the oligomeric state, the secondary structure, and the thermal stability of TF in the absence and presence of Zn.

Ca Regulates ERp57-Calnexin Complex Formation.

ERp57, a member of the protein disulfide isomerase family, is a ubiquitous disulfide catalyst that functions in the oxidative folding of various clients in the mammalian endoplasmic reticulum (ER). In concert with ER lectin-like chaperones calnexin and calreticulin (CNX/CRT), ERp57 functions in virtually all folding stages from co-translation to post-translation, and thus plays a critical role in maintaining protein homeostasis, with direct implication for pathology. Here, we present mechanisms by which Ca regulates the formation of the ERp57-calnexin complex.

A unique leucine-valine adhesive motif supports structure and function of protein disulfide isomerase P5 via dimerization.

P5, also known as PDIA6, is a PDI family member involved in the ER quality control. Here, we revealed that P5 dimerizes via a unique adhesive motif contained in the N-terminal thioredoxin-like domain. Unlike conventional leucine zipper motifs with leucine residues every two helical turns on ∼30-residue parallel α helices, this adhesive motif includes periodic repeats of leucine/valine residues at the third or fourth position spanning five helical turns on 15-residue anti-parallel α helices.

Conjugate of Thiol and Guanidyl Units with Oligoethylene Glycol Linkage for Manipulation of Oxidative Protein Folding.

Oxidative protein folding is a biological process to obtain a native conformation of a protein through disulfide-bond formation between cysteine residues. In a cell, disulfide-catalysts such as protein disulfide isomerase promote the oxidative protein folding. Inspired by the active sites of the disulfide-catalysts, synthetic redox-active thiol compounds have been developed, which have shown significant promotion of the folding processes.

PDI Family Members as Guides for Client Folding and Assembly.

Complicated and sophisticated protein homeostasis (proteostasis) networks in the endoplasmic reticulum (ER), comprising disulfide catalysts, molecular chaperones, and their regulators, help to maintain cell viability. Newly synthesized proteins inserted into the ER need to fold and assemble into unique native structures to fulfill their physiological functions, and this is assisted by protein disulfide isomerase (PDI) family. Herein, we focus on recent advances in understanding the detailed mechanisms of PDI family members as guides for client folding and assembly to ensure the efficient production of secretory proteins.

A novel soybean protein disulphide isomerase family protein possesses dithiol oxidation activity: identification and characterization of GmPDIL6.

Secretory and membrane proteins synthesized in the endoplasmic reticulum (ER) are folded with intramolecular disulphide bonds, viz. oxidative folding, catalysed by the protein disulphide isomerase (PDI) family proteins. Here, we identified a novel soybean PDI family protein, GmPDIL6.

Regulation of plant ER oxidoreductin 1 (ERO1) activity for efficient oxidative protein folding.

In the endoplasmic reticulum (ER), ER oxidoreductin 1 (ERO1) catalyzes intramolecular disulfide-bond formation within its substrates in coordination with protein-disulfide isomerase (PDI) and related enzymes. However, the molecular mechanisms that regulate the ERO1-PDI system in plants are unknown. Reduction of the regulatory disulfide bonds of the ERO1 from soybean, GmERO1a, is catalyzed by enzymes in five classes of PDI family proteins.

The Protein Disulfide Isomerase Family: from proteostasis to pathogenesis.

In mammalian cells, nearly one-third of proteins are inserted into the endoplasmic reticulum (ER), where they undergo oxidative folding and chaperoning assisted by approximately 20 members of the protein disulfide isomerase family (PDIs). PDIs consist of multiple thioredoxin-like domains and recognize a wide variety of proteins via highly conserved interdomain flexibility. Although PDIs have been studied intensely for almost 50 years, exactly how they maintain protein homeostasis in the ER remains unknown, and is important not only for fundamental biological understanding but also for protein misfolding- and aggregation-related pathophysiology.

Coupling effects of thiol and urea-type groups for promotion of oxidative protein folding.

Coupling of thiol and urea-type -NHC([double bond, length as m-dash]X)NH2 (X = O or NH) groups is effective in promoting oxidative protein folding. In particular, a thiol compound coupled with a guanidyl (X = NH) group significantly accelerates the rates of folding processes and enhances the yields of native proteins.

Identification and characterization of GmPDIL7, a soybean ER membrane-bound protein disulfide isomerase family protein.

Unlabelled: Most proteins synthesized in the endoplasmic reticulum (ER) possess intramolecular and intermolecular disulfide bonds, which play an important role in the conformational stability and function of proteins. Hence, eukaryotic cells contain protein disulfide bond formation pathways such as the protein disulfide isomerase (PDI)-ER oxidoreductin 1 (Ero1) system in the ER lumen. In this study, we identified soybean PDIL7 (GmPDIL7), a novel soybean ER membrane-bound PDI family protein, and determined its enzymatic properties.

Cooperative Protein Folding by Two Protein Thiol Disulfide Oxidoreductases and 1 in Soybean.

Most proteins produced in the endoplasmic reticulum (ER) of eukaryotic cells fold via disulfide formation (oxidative folding). Oxidative folding is catalyzed by protein disulfide isomerase (PDI) and PDI-related ER protein thiol disulfide oxidoreductases (ER oxidoreductases). In yeast and mammals, ER oxidoreductin-1s (Ero1s) supply oxidizing equivalent to the active centers of PDI.

Disulfide bond formation activity of soybean quiescin sulfhydryl oxidase.

Unlabelled: Multiple enzymatic systems can catalyse protein disulfide bond formation in the endoplasmic reticulum (ER) of eukaryotic cells. The enzyme quiescin sulfhydryl oxidase (QSOX) catalyses disulfide bond formation in unfolded proteins via the reduction of oxygen. We found two QSOX homologues in the soybean genome database, Glycine max QSOX (GmQSOX)1 and GmQSOX2, which encode proteins composed of an N-terminal signal peptide, a thioredoxin-like domain, an FAD-binding domain, Erv/ALR, and a transmembrane region near the C terminus.

Accumulation of β-conglycinin in soybean cotyledon through the formation of disulfide bonds between α'- and α-subunits.

β-Conglycinin, one of the major soybean (Glycine max) seed storage proteins, is folded and assembled into trimers in the endoplasmic reticulum and accumulated into protein storage vacuoles. Prior experiments have used soybean β-conglycinin extracted using a reducing buffer containing a sulfhydryl reductant such as 2-mercaptoethanol, which reduces both intermolecular and intramolecular disulfide bonds within the proteins. In this study, soybean proteins were extracted from the cotyledons of immature seeds or dry beans under nonreducing conditions to prevent the oxidation of thiol groups and the reduction or exchange of disulfide bonds.