Catalysis of Protein Folding by Protein Disulfide Isomerase and Small-Molecule Mimics

Kersteen, Elizabeth A.; Raines, Ronald T.

doi:10.1089/152308603768295159

Cited by 66 publications

(45 citation statements)

References 104 publications

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“…The pro-sequence of BPTI contains a single cysteine residue that is required for this rate enhancement. Further, a single cysteine residue tethered to the C-terminus of mature BPTI through an artificial (SerGly-Gly) 3 linker provides rate and yield enhancements similar to those seen with pro-BPTI. Although the pro-sequence is not necessarily important for the physiological folding of BPTI (72), these results suggest a general strategy for more efficient protein folding.…”

Section: Prospectusmentioning

confidence: 97%

The CXC Motif: A Functional Mimic of Protein Disulfide Isomerase

Woycechowsky

Raines

2003

Biochemistry

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Protein disulfide isomerase (PDI) utilizes the active-site sequence Cys-Gly-His-Cys (CGHC; E°′ = −180 mV) to effect thiol-disulfide interchange during oxidative protein folding. Here, the Cys-GlyCys-NH 2 (CGC) peptide is shown to have a disulfide reduction potential (E°′ = −167 mV) that is close to that of PDI. This peptide has a thiol acid dissociation constant (pK a = 8.7) that is lower than that of glutathione. These attributes endow the CGC peptide with substantial disulfide isomerization activity. Escherichia coli thioredoxin (Trx) utilizes the active-site sequence Cys-Gly-Pro-Cys (CGPC; E°′ = −270 mV) to effect disulfide reduction. Removal of the proline residue from the Trx active site yields a CGC active site with a greatly destabilized disulfide bond (E°′ ≥ −200 mV). The ΔP34 variant retains high conformational stability and remains a substrate for thioredoxin reductase. In contrast to the reduced form of the wild-type enzyme, the reduced form of ΔP34 Trx has disulfide isomerization activity, which is 25-fold greater than that of the CGC peptide. Thus, the rational deletion of an active-site residue can bestow a new and desirable function upon an enzyme. Moreover, the CGC motif, in both a peptide and a protein, provides functional mimicry of PDI.Protein disulfide isomerase (PDI 1 ; EC 5.3.4.1 (1-3)) is the most efficient known catalyst of oxidative protein folding (4). A thiol-disulfide oxidoreductase with an archetypal active-site motif: Cys-Xaa-Xaa-Cys (CXXC, where X is any amino acid), PDI catalyzes the formation, reduction, and isomerization of disulfide bonds in a protein substrate (5). The product contains the most stable arrangement of disulfide bonds in a particular redox environment (6).Oxidoreductases with CXXC motifs vary in their disulfide bond stability, subcellular localization, and biological function (7-9). For example, PDI, a resident of the endoplasmic reticulum, has a CGHC active site with E°′ = −180 mV (10), and is essential for catalysis of disulfide isomerization in Saccharomyces cerevisiae (5). Escherichia coli thioredoxin (Trx) has a CGPC active site (11) with E°′ = −270 mV (12), and functions as a cytosolic reductant (13). DsbA has a CPHC active site with E°′ = −122 mV (14), and catalyzes the oxidation of proteins that have been secreted to the E. coli periplasm (15). These three enzymes are believed † This work was supported by grant BES04563 (NSF). K.J.W. was supported by a WARF predoctoral fellowship and Chemistry-Biology Interface Training Grant GM08505 (NIH).*To whom all correspondence should be addressed at the Department of Biochemistry, University of Wisconsin-Madison, 433 Babcock Drive, Madison, WI 53706-1544. Telephone: (608) . raines@biochem.wisc.edu. ‡ Present address: Laboratorium für Organische Chemie, Swiss Federal Institute of Technology, ETH-Hönggerberg, CH-8093 Zürich, Switzerland 1 BMC, (±)-trans-1,2-bis(mercaptoacetamido)cyclohexane; BPTI, bovine pancreatic trypsin inhibitor; DTNB, 5,5′-dithiobis(2-nitrobenzoic acid); DTT, dithiothreitol; EDTA, e...

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Section: Prospectusmentioning

confidence: 97%

The CXC Motif: A Functional Mimic of Protein Disulfide Isomerase

Woycechowsky

Raines

2003

Biochemistry

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“…The process of native disulfide formation involves both the oxidation of dithiols to disulfides as well as the rearrangement of non-native to native disulfide bonds. PDI is involved in both these steps (for a review of the mechanisms, see Kersteen and Raines 2003). Until recently, the source of oxidizing equivalents necessary for these reactions remained elusive, until it was found that in S. cerevisiae the native ER protein Ero1 transfers disulphide bonds to PDI (Frand and Kaiser 1998;Pollard et al 1998; for review, see Frand et al 2000).…”

Section: Cellular Redox Homeostasis and Organellesmentioning

confidence: 99%

Oxygen free radicals and redox biology of organelles

Moldovan

2004

Histochem Cell Biol

166

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The presence and supposed roles of reactive oxygen species (ROS) were reported in literature in a myriad of instances. However, the breadth and depth of their involvement in cellular physiology and pathology, as well as their relationship to the redox environment can only be guessed from specialized reports. Whatever their circumstances of formation or consequences, ROS seem to be conspicuous components of intracellular milieu. We sought to verify this assertion, by collecting the available evidence derived from the most recent publications in the biomedical field. Unlike other reviews with similar objectives, we centered our analysis on the subcellular compartments, namely on organelles, grouped according to their major functions. Thus, plasma membrane is a major source of ROS through NAD(P)H oxidases located on either side. Enzymes of the same class displaying low activity, as well as their components, are also present free in cytoplasm, regulating the actin cytoskeleton and cell motility. Mitochondria can be a major source of ROS, mainly in processes leading to apoptosis. The protein synthetic pathway (endoplasmic reticulum and Golgi apparatus), including the nucleus, as well as protein turnover, are all exquisitely sensitive to ROS-related redox conditions. The same applies to the degradation pathways represented by lysosomes and peroxisomes. Therefore, ROS cannot be perceived anymore as a mere harmful consequence of external factors, or byproducts of altered cellular metabolism. This may explain why the indiscriminate use of anti-oxidants did not produce the expected "beneficial" results in many medical applications attempted so far, underlying the need for a deeper apprehension of the biological roles of ROS, particularly in the context of the higher cellular order of organelles.

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“…PDI is a eukaryotic disulfide isomerase, having four thioredoxin domains (Kersteen and Raines 2003). DsbC is a periplasmic disulfide isomerase of Gram-negative bacteria and displays about 30% of the activities of eukaryotic PDI as isomerase and as thiol-protein oxidoreductase (Chen et al 1999).…”

Section: Discussionmentioning

confidence: 99%

MTH1745, a protein disulfide isomerase-like protein from thermophilic archaea, Methanothermobacter thermoautotrophicum involving in stress response

Ding

Zhao

et al. 2008

Cell Stress and Chaperones

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MTH1745 is a putative protein disulfide isomerase characterized with 151 amino acid residues and a CPAC active-site from the anaerobic archaea Methanothermobacter thermoautotrophicum. The potential functions of MTH1745 are not clear. In the present study, we show a crucial role of MTH1745 in protecting cells against stress which may be related to its functions as a disulfide isomerase and its chaperone properties. Using real-time polymerase chain reaction analyses, the level of MTH1745 messenger RNA (mRNA) in the thermophilic archaea M. thermoautotrophicum was found to be stress-induced in that it was significantly higher under low (50°C) and high (70°C) growth temperatures than under the optimal growth temperature for the organism (65°C). Additionally, the expression of MTH1745 mRNA was up-regulated by cold shock (4°C). Furthermore, the survival of MTH1745 expressing Escherichia coli cells was markedly higher than that of control cells in response to heat shock (51.0°C). These results indicated that MTH1745 plays an important role in the resistance of stress. By assay of enzyme activities in vitro, MTH1745 also exhibited a chaperone function by promoting the functional folding of citrate synthase after thermodenaturation. On the other hand, MTH1745 was also shown to function as a disulfide isomerase on the refolding of denatured and reduced ribonuclease A. On the basis of its single thioredoxin domain, function as a disulfide isomerase, and its chaperone activity, we suggest that MTH1745 may be an ancient protein disulfide isomerase. These studies may provide clues to the understanding of the function of protein disulfide isomerase in archaea.

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Catalysis of Protein Folding by Protein Disulfide Isomerase and Small-Molecule Mimics

Cited by 66 publications

References 104 publications

The CXC Motif: A Functional Mimic of Protein Disulfide Isomerase

The CXC Motif: A Functional Mimic of Protein Disulfide Isomerase

Oxygen free radicals and redox biology of organelles

MTH1745, a protein disulfide isomerase-like protein from thermophilic archaea, Methanothermobacter thermoautotrophicum involving in stress response

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