An Amphiphilic Selenide Catalyst Behaves Like a Hybrid Mimic of Protein Disulfide Isomerase and Glutathione Peroxidase 7

Abstract

Protein disulfide isomerase (PDI) and glutathione peroxidase 7 (GPx7) cooperatively promote the oxidative folding of disulfide (SS)-containing proteins in endoplasmic reticulum by recognizing the nascent proteins to convert them into the native folds by means of SS formation and SS isomerization and by catalyzing reoxidation of reduced PDI with H2O2, respectively. In this study, new amphiphilic selenides with a long-chain alkyl group were designed as hybrid mimics of PDI and GPx7 and were applied to the refolding of reduced hen egg-white lysozyme (HEL-R). Competitive SS formation at pH 4 using HEL-R and glutathione (GSH) in the presence of the selenide catalyst and H2O2 showed that the amphiphilic selenides can preferentially catalyze SS formation of HEL-R, probably on account of hydrophobic interactions between the protein and the catalyst. In contrast, simple water-soluble selenides did not exhibit such behavior. In addition, when the pH of the solution was adjusted to 8.5 after the SS formation, surviving GSH promoted the SS isomerization of misfolded HEL to recover the native SS linkages. Thus, the amphiphilic selenides designed here could mimic the function of the PDI-GPx7 system. The combination of a water-soluble selenide and a long-chain alkyl group would be a useful motif in designing medicines for both protein misfolding diseases and antioxidant therapy.

Glutathione Peroxidase-Like Activity of Amino-Substituted Water-Soluble Cyclic Selenides: A Shift of the Major Catalytic Cycle in Methanol

“…Compounds 5 and 6 were synthesized by applying a similar protocol reported previously [ 33 ] ( Scheme 2 ). Boc-protected mesylates 12a and 12b were first prepared from diethyl aminomalonate hydrochloride ( 10 ) or l -glutamic acid ( 11 ), respectively, by following previous methods with slight modifications [ 34 , 35 ].…”

Protein Folding in the Presence of Water‐Soluble Cyclic Diselenides with Novel Oxidoreductase and Isomerase Activities

“…The thioredoxin‐like domains containing cysteine residues (CXXC motifs) at PDI active sites inspired the development of disulfide‐ and diselenide‐based compounds as catalysts for in vitro oxidative protein folding and/or refolding (reactions A and/or D) . We recently reported that long‐chain, alkyl‐group‐conjugated cyclic monoselenides can recognize a reduced protein and catalytically introduce SS bonds randomly in a protein in the presence of H 2 O 2 (reaction B) . Thus, these PDI mimics mediated one or more out of reactions A, B, and D. Much less attention, however, has been directed towards SS reduction (reaction C).…”

Basic Amino Acid Conjugates of 1,2‐Diselenan‐4‐amine with Protein Disulfide Isomerase‐like Functions as a Manipulator of Protein Quality Control

“…The recovered enzymatic activity of HEL in the solution was estimated by the activity assay for HEL as described previously. [20]…”

“…After centrifugation of the sample solution (14,000 rpm, for 3 min), the recovered enzymatic activity of HEL was estimated by the activity assay for HEL as described previously. [20] Oxidative folding of HEL at a high protein concentration R HEL solution (200 μM) dissolved in 100 mM Tris-HCl buffer solution containing 1 mM EDTA and 4 M urea at pH 7.5 were prepared as described above. The R HEL solution (50 μL) was mixed with 2.67 mM GSH (75 μL) and 0 or 533 μM diselenide compound or GSSG (75 μL), which were prepared by the solution without urea, and the resulting mixture was incubated at 37.0°C.…”

“…The recovered enzymatic activity of HEL was estimated by the activity assay for HEL as described previously. [20] The experiment for each compound was performed in parallel and repeated four times.…”