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Silvennoinen, Laura; Karvonen, P.; Koivunen, Peppi; Myllyharju, Johanna; Kivirikko, Kari I.; Kilpeläinen, Ilkka

doi:10.1023/a:1011291316810

Cited by 8 publications

(9 citation statements)

References 7 publications

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“…Expression constructs for ERp1-123 and ERp1-119, containing NcoI and HindIII sites at their 5Ј and 3Ј ends, were generated by PCR and cloned into the corresponding sites in pET32a (Novagen). ERp349 -468 was generated as described earlier (27).…”

Section: Construction Of Escherichia Coli Expression Vectors For Erp5mentioning

confidence: 99%

“…Fractions containing the protein were pooled and concentrated, and the protein concentrations were determined as above. ERp349 -468 was produced and purified as described earlier (27).…”

Section: Construction Of Escherichia Coli Expression Vectors For Erp5mentioning

confidence: 99%

“…The digestions with proteinase K resulted in the largest population of bands. The N termini of the 16.5-and 14.5-kDa (25); line E, location of the verified secondary structure elements of ERp57, domains a 2 and aЈ from a chemical shift index derived from C␣ and H␣ chemical shifts (27); lines 1 and 2, predictions of secondary structure elements of ERp57 by two programs, PredictProtein and ProfPrediction, available at cubic.bioc.columbia.edu/pp/ and www.aber.ac.uk/ϳphiwww/prof/, respectively. fragments were sequenced, and both were found to begin with the histidine tag and were likely to extend to about residues 140 and 120, respectively (Fig.…”

Section: Gel Filtration Reveals That Erp57 Expressed In Insect Cellsmentioning

confidence: 99%

“…No three-dimensional structure is available for ERp57, but nuclear magnetic resonance (NMR) characterization of its aЈ domain has shown that it is structurally related to the corresponding PDI domain and also possesses the thioredoxin fold (27).…”

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confidence: 99%

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Identification and Characterization of Structural Domains of Human ERp57

Silvennoinen

Myllyharju

Ruoppolo

et al. 2004

Journal of Biological Chemistry

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The amino acid sequence of ERp57, which functions in the endoplasmic reticulum together with the lectins calreticulin and calnexin to achieve folding of newly synthesized glycoproteins, is highly similar to that of protein disulfide isomerase (PDI), but they have their own distinct roles in protein folding. We have characterized the domain structure of ERp57 by limited proteolysis and N-terminal sequencing and have found it to be similar but not identical to that of PDI. ERp57 had three major protease-sensitive regions, the first of which was located between residues 120 and 150, the second between 201 and 215, and the third between 313 and 341, the data thus being consistent with a four-domain structure abba. Recombinant expression in Escherichia coli was used to verify the domain boundaries. Each single domain and a ba double domain could be produced in the form of soluble, folded polypeptides, as verified by circular dichroism spectra and urea gradient gel electrophoresis. When the ability of ERp57 and its a and a domains to fold denatured RNase A was studied by electrospray mass analyses, ERp57 markedly enhanced the folding rate at early time points, although less effectively than PDI, but was an ineffective catalyst of the overall process. The a and a domains produced only minor, if any, increases in the folding rate at the early stages and no increase at the late stages. Interaction of the soluble ERp57 domains with the P domain of calreticulin was studied by chemical cross-linking in vitro. None of the single ERp57 domains nor the ba double domain could be cross-linked to the P domain, whereas cross-linking was obtained with a hybrid ERpabbPDIac polypeptide but not with ERpabPDIbac, indicating that multiple domains are involved in this protein-protein interaction and that the b domain of ERp57 cannot be replaced by that of PDI.ERp57 is a protein resident in the endoplasmic reticulum (ER) 1 that functions as a co-chaperone in glycoprotein folding with the lectins calnexin (CNX) and its soluble homologue calreticulin (CRT) (for reviews, see Refs. 1 and 2). ERp57 is a member of the protein disulfide isomerase (PDI) oxidoreductase family (3, 4) and has two thioredoxin-like domains with -Cys-Gly-His-Cys-catalytic site motifs in positions corresponding to the thioredoxin-like domains a and aЈ of PDI, which consists altogether of domains a, b, bЈ, and aЈ plus an acidic C-terminal extension c (5). ERp57 also shows a significant sequence similarity to PDI in regions corresponding to domains b and bЈ (6). No similarity is found in the organization of the human genes or gene loci for ERp57 and PDI, however (7). Despite the 56% overall amino acid sequence similarity between PDI and ERp57, the latter does not substitute for PDI as the ␤ subunit in the assembly of the collagen prolyl 4-hydroxylase ␣ 2 ␤ 2 tetramer (6), an enzyme involved in the synthesis of collagens (8 -10), but the N-terminal domains a and b of ERp57 can in part substitute for those of PDI in this assembly (11). ERp57 possesses disulfide oxidoreduc...

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Section: Construction Of Escherichia Coli Expression Vectors For Erp5mentioning

confidence: 99%

“…Fractions containing the protein were pooled and concentrated, and the protein concentrations were determined as above. ERp349 -468 was produced and purified as described earlier (27).…”

Section: Construction Of Escherichia Coli Expression Vectors For Erp5mentioning

confidence: 99%

Section: Gel Filtration Reveals That Erp57 Expressed In Insect Cellsmentioning

confidence: 99%

mentioning

confidence: 99%

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Identification and Characterization of Structural Domains of Human ERp57

Silvennoinen

Myllyharju

Ruoppolo

et al. 2004

Journal of Biological Chemistry

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“…For ERp57, the information regarding the three-dimensional structure is limited to NMR assignments of the a domain (31), and like PDI, ERp57 contains two CGHC active-site sequence motifs. Overall, the protein remains poorly characterized at the molecular level.…”

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confidence: 99%

ERp57 Is a Multifunctional Thiol-Disulfide Oxidoreductase

Frickel

Frei

Bouvier

et al. 2004

Journal of Biological Chemistry

179

129

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The thiol-disulfide oxidoreductase ERp57 is a soluble protein of the endoplasmic reticulum and the closest known homologue of protein disulfide isomerase. The protein interacts with the two lectin chaperones calnexin and calreticulin and thereby promotes the oxidative folding of newly synthesized glycoproteins. Here we have characterized several fundamental structural and functional properties of ERp57 in vitro, such as the domain organization, shape, redox potential, and the ability to catalyze different thiol-disulfide exchange reactions. Like protein disulfide isomerase, we find ERp57 to be comprised of four structural domains. The protein has an elongated shape of 3.4 ؎ 0.1 nm in diameter and 16.8 ؎ 0.5 nm in length. The two redox-active a and a domains were determined to have redox potentials of ؊0.167 and ؊0.156 V, respectively. Furthermore, ERp57 was shown to efficiently catalyze disulfide reduction, disulfide isomerization, and dithiol oxidation in substrate proteins. The implications of these findings for the function of the protein in vivo are discussed.

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Multiple catalytically active thioredoxin folds: a winning strategy for many functions

Pedone

Limauro

D’Ambrosio

et al. 2010

Cell. Mol. Life Sci.

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The Thioredoxin (Trx) fold is a versatile protein scaffold consisting of a four-stranded β-sheet surrounded by three α-helices. Various insertions are possible on this structural theme originating different proteins, which show a variety of functions and specificities. During evolution, the assembly of different Trx fold domains has been used many times to build new multi-domain proteins able to perform a large number of catalytic functions. To clarify the interaction mode of the different Trx domains within a multi-domain structure and how their combination can affect catalytic performances, in this review, we report on a structural and functional analysis of the most representative proteins containing more than one catalytically active Trx domain: the eukaryotic protein disulfide isomerases (PDIs), the thermophilic protein disulfide oxidoreductases (PDOs) and the hybrid peroxiredoxins (Prxs).

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Cited by 8 publications

References 7 publications

Identification and Characterization of Structural Domains of Human ERp57

Identification and Characterization of Structural Domains of Human ERp57

ERp57 Is a Multifunctional Thiol-Disulfide Oxidoreductase

Multiple catalytically active thioredoxin folds: a winning strategy for many functions

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