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• W2077047894 abstract "p58/ERGIC-53 is an animal calcium-dependent lectin that cycles between the endoplasmic reticulum (ER) and the Golgi complex and appears to act as a cargo receptor for a subset of soluble glycoproteins exported from the ER. We have determined the crystal structure of the carbohydrate recognition domain (CRD) of p58, the rat homologue of human ERGIC-53, to 1.46 Å resolution. The fold and ligand binding site are most similar to those of leguminous lectins. The structure also resembles that of the CRD of the ER folding chaperone calnexin and the neurexins, a family of non-lectin proteins expressed on neurons. The CRD comprises one concave and one convex β-sheet packed into a β-sandwich. The ligand binding site resides in a negatively charged cleft formed by conserved residues. A large surface patch of conserved residues with a putative role in protein-protein interactions and oligomerization lies on the opposite side of the ligand binding site. Together with previous functional data, the structure defines a new and expanding class of calcium-dependent animal lectins and provides a starting point for the understanding of glycoprotein sorting between the ER and the Golgi. p58/ERGIC-53 is an animal calcium-dependent lectin that cycles between the endoplasmic reticulum (ER) and the Golgi complex and appears to act as a cargo receptor for a subset of soluble glycoproteins exported from the ER. We have determined the crystal structure of the carbohydrate recognition domain (CRD) of p58, the rat homologue of human ERGIC-53, to 1.46 Å resolution. The fold and ligand binding site are most similar to those of leguminous lectins. The structure also resembles that of the CRD of the ER folding chaperone calnexin and the neurexins, a family of non-lectin proteins expressed on neurons. The CRD comprises one concave and one convex β-sheet packed into a β-sandwich. The ligand binding site resides in a negatively charged cleft formed by conserved residues. A large surface patch of conserved residues with a putative role in protein-protein interactions and oligomerization lies on the opposite side of the ligand binding site. Together with previous functional data, the structure defines a new and expanding class of calcium-dependent animal lectins and provides a starting point for the understanding of glycoprotein sorting between the ER and the Golgi. endoplasmic reticulum carbohydrate recognition domain Protein Data Bank Secretory and membrane proteins undergo a quality control process that assures their proper folding, oligomerization, and maturation before exit from the endoplasmic reticulum (ER)1 (1Ellgaard L. Molinari M. Helenius A. Science. 1999; 286: 1882-1888Crossref PubMed Scopus (1064) Google Scholar). This is followed by the critical step of selection of cargo proteins to be exported from the ER to the Golgi complex via the ERGIC (ER-Golgi intermediate compartment). Export is then mediated by COPII vesicles (2Klumperman J. Curr. Opin. Cell Biol. 2000; 12: 445-449Crossref PubMed Scopus (110) Google Scholar). Although many proteins may become incorporated into COPII vesicles by default (bulk-flow transport), it is now generally believed that export is an active regulated process, whereby cargo molecules are first localized to ER exit sites and then selectively incorporated into COPII vesicles by one of several mechanisms (2Klumperman J. Curr. Opin. Cell Biol. 2000; 12: 445-449Crossref PubMed Scopus (110) Google Scholar, 3Balch W.E. McCaffery J.M. Plutner H. Farquhar M.G. Cell. 1994; 76: 841-852Abstract Full Text PDF PubMed Scopus (333) Google Scholar). Selective export of soluble proteins may occur by interaction with export receptors harboring motifs recognized by COPII coatomers. The p58 and ERGIC-53/MR60 proteins are the most commonly used markers for the ERGIC (4Saraste J. Palade G.E. Farquhar M.G. J. Cell Biol. 1987; 105: 2021-2029Crossref PubMed Scopus (125) Google Scholar, 5Schweizer A. Fransen J.A.M. Bächi T. Ginsel L. Hauri H.P. J. Cell Biol. 1988; 107: 1643-1653Crossref PubMed Scopus (375) Google Scholar). p58 (4Saraste J. Palade G.E. Farquhar M.G. J. Cell Biol. 1987; 105: 2021-2029Crossref PubMed Scopus (125) Google Scholar) and ERGIC-53 (5Schweizer A. Fransen J.A.M. Bächi T. Ginsel L. Hauri H.P. J. Cell Biol. 1988; 107: 1643-1653Crossref PubMed Scopus (375) Google Scholar) were originally identified as proteins reacting with antibodies prepared against Golgi membrane fractions, whereas MR60 was identified as a mannose-binding protein (6Pimpaneau V. Midoux P. Monsigny M. Roche A.C. Carbohydr. Res. 1991; 213: 95-108Crossref PubMed Scopus (24) Google Scholar). Subsequent cDNA cloning showed that ERGIC-53 (7Schindler R. Itin C. Zerial M. Lottspeich F. Hauri H.-P. Eur. J. Cell Biol. 1993; 61: 1-9PubMed Google Scholar) and MR60 (8Arar C. Le Carpentier V. Caer J.-P. Monsigny M. Legrand A. Roche A.-C. J. Biol. Chem. 1995; 270: 3551-3553Abstract Full Text Full Text PDF PubMed Scopus (102) Google Scholar) were identical to each other and represented the human homologue of the rat p58 protein (9Lahtinen U. Hellman U. Wernstedt C. Saraste J. Pettersson R. J. Biol. Chem. 1996; 271: 4031-4037Abstract Full Text Full Text PDF PubMed Scopus (78) Google Scholar). p58/ERGIC-53/MR60 is a type I transmembrane, nonglycosylated protein with a lumenal domain, a transmembrane domain, and a short cytoplasmic domain. In cells, it is present as dimers and hexamers (9Lahtinen U. Hellman U. Wernstedt C. Saraste J. Pettersson R. J. Biol. Chem. 1996; 271: 4031-4037Abstract Full Text Full Text PDF PubMed Scopus (78) Google Scholar, 10Lahtinen U. Svensson K. Pettersson R.F. Eur. J. Biochem. 1999; 260: 392-397Crossref PubMed Scopus (21) Google Scholar). The lumenal domain can be divided into two subdomains, an N-terminal carbohydrate recognition domain (CRD) (residues 31–285) and a membrane-proximal α-helical coiled domain (residues 290–460) (10Lahtinen U. Svensson K. Pettersson R.F. Eur. J. Biochem. 1999; 260: 392-397Crossref PubMed Scopus (21) Google Scholar). The cytoplasmic tail contains a KKFF sequence at its extreme C terminus that is essential for both ER exit and Golgi retrieval (11Kappeler F. Klopfenstein D.R.Ch. Foguet M. Paccaud J.-P. Hauri H.-P. J. Biol. Chem. 1997; 272: 31802-31808Abstract Full Text Full Text PDF Scopus (226) Google Scholar), meaning that p58/ERGIC-53/MR60 cycles between the ER and Golgi compartment (12Klumperman J. Schweizer A. Clausen H. Tang B.L. Hong W. Oorschot V. Hauri H.-P. J. Cell Sci. 1998; 111: 3411-3425PubMed Google Scholar). The identification of MR60 as a mannose-binding protein (8Arar C. Le Carpentier V. Caer J.-P. Monsigny M. Legrand A. Roche A.-C. J. Biol. Chem. 1995; 270: 3551-3553Abstract Full Text Full Text PDF PubMed Scopus (102) Google Scholar) and mutagenesis studies of ERGIC-53, in which the calcium-dependent binding of mannose to this protein was abolished (13Itin C. Roche A.-C. Monsigny M. Hauri H.-P. Mol. Biol. Cell. 1996; 7: 483-493Crossref PubMed Scopus (154) Google Scholar), support the suggestion that p58/ERGIC-53 serves as a mammalian intracellular lectin (14Fiedler K. Simons K. Cell. 1994; 77: 625-626Abstract Full Text PDF PubMed Scopus (133) Google Scholar). This proposition was also based on the similarity between ERGIC-53 and VIP-36, a membrane protein isolated from Madin-Darby canine kidney cells (15Fiedler K. Parton R.G. Kellner R. Etzold T. Simons K. EMBO J. 1994; 13: 1729-1740Crossref PubMed Scopus (193) Google Scholar), shown to bind mannose, and localized in the early secretory pathway (16Fullerkrug J. Scheiffele P. Simons K. J. Cell Sci. 1999; 112: 2813-2821PubMed Google Scholar, 17Hara-Kuge S. Ohkura T. Seko A. Yamashita K. Glycobiology. 1999; 9: 833-839Crossref PubMed Scopus (69) Google Scholar). It was proposed that p58/ERGIC-53 and VIP-36 constituted a new class of animal lectins (14Fiedler K. Simons K. Cell. 1994; 77: 625-626Abstract Full Text PDF PubMed Scopus (133) Google Scholar). Blocking the export of ERGIC-53 from the ER impaired but did not block export of the lysosomal enzyme cathepsin C and cathepsin Z-related protein (18Vollenweider F. Kappeler F. Itin C. Hauri H.-P. J. Cell Biol. 1998; 142: 377-389Crossref PubMed Scopus (134) Google Scholar). Furthermore, ERGIC-53 could be cross-linked to a cathepsin Z-related protein. The glycan structure binding to ERGIC-53 in the ER was suggested to be a nine-mannose form (Man9),i.e. the asparagine-linked core glycan from which three terminal glucose residues have been trimmed (19Appenzeller C. Andersson H. Kappeler F. Hauri H.-P. Nat. Cell Biol. 1999; 1: 330-334Crossref PubMed Scopus (273) Google Scholar). Efficient secretion of coagulation factors V and VIII requires a functional ERGIC-53 (20Moussalli M. Pipe S.W. Hauri H.-P. Nichols W.C. Ginsburg D. Kaufman R.J. J. Biol. Chem. 1999; 274: 32539-32542Abstract Full Text Full Text PDF PubMed Scopus (117) Google Scholar), and mutations in the gene coding for ERGIC-53 cause a rare hereditary bleeding disorder, a combined deficiency of factors V and VIII (21Nichols W.C. Seligsohn U. Zivelin A. Terry V.H. Hertel C.E. Weatley M.A. Moussalli M. Hauri H.P. Ciavarella N. Kaufman R.J. Ginsburg D. Cell. 1998; 93: 61-70Abstract Full Text Full Text PDF PubMed Scopus (345) Google Scholar). Taken together, these data strongly suggest that p58/ERGIC-53/MR60 functions as a lectin-like receptor involved in facilitating export from the ER of a subset of secretory glycoproteins (22Hauri H.-P. Kappeler F. Andersson H. Appenzeller C. J. Cell Sci. 2000; 113: 587-596Crossref PubMed Google Scholar). As a first step toward a better understanding of how glycans on cargo molecules interact with the lectin receptor, we have determined the crystal structure of the CRD of p58. The structure reveals a link between leguminous and animal lectins and defines a new class of animal calcium-dependent lectins that function in the secretory pathway. The CRD of rat p58/ERGIC-53 (residues 31–285) was defined by a combination of sequence alignment, secondary structure prediction, and limited proteolysis. The CRD (including the N-terminal signal sequence comprising residues 1–30 and a 6xHis tag) was produced in insect cells using a baculovirus vector and purified as described elsewhere (23Velloso L.M. Svensson K. Lahtinen U. Schneider G. Pettersson R.F. Linqvist Y. Acta Crystallogr. Sect. D Biol. Crystallogr. 2002; 58: 536-538Crossref PubMed Scopus (8) Google Scholar). The region encompassing residues 286–478, which is not present in the construct whose structure is described here, is the oligomerization domain of p58/ERGIC-53 and is required for dimerization and formation of hexamers (10Lahtinen U. Svensson K. Pettersson R.F. Eur. J. Biochem. 1999; 260: 392-397Crossref PubMed Scopus (21) Google Scholar). Therefore, the CRD is monomeric in solution as assayed by native gel electrophoresis and gel filtration chromatography (data not shown). Crystals were grown by vapor diffusion from hanging drops containing equal volumes of a 10 mg/ml protein solution in 10 mm Tris-HCl, pH 7.5, 1 mmCaCl2, and well solution. The drops were equilibrated against 1 ml of well solution that consisted of 100 mmNa-HEPES, pH 7.25, 1.6 m Li2SO4, and 10 mm EDTA, as described previously (23Velloso L.M. Svensson K. Lahtinen U. Schneider G. Pettersson R.F. Linqvist Y. Acta Crystallogr. Sect. D Biol. Crystallogr. 2002; 58: 536-538Crossref PubMed Scopus (8) Google Scholar). The crystals belong to the orthorhombic space group I222, with cell dimensionsa = 49.6 Å, b = 86.1 Å, andc = 128.1 Å, and they have one monomer in the asymmetric unit (23Velloso L.M. Svensson K. Lahtinen U. Schneider G. Pettersson R.F. Linqvist Y. Acta Crystallogr. Sect. D Biol. Crystallogr. 2002; 58: 536-538Crossref PubMed Scopus (8) Google Scholar). X-ray data used for the structure determination were collected on a MAR 300mm Image plate detector mounted on a Rigaku R200 x-ray generator, operating at 50 kV and 90 mA at 110 K. Crystals were transferred to a solution containing 1.2m Li2SO4, 0.1 mNa-HEPES, pH 7.25, and 20% PEG400, allowed to equilibrate for a few seconds, and frozen in a stream of nitrogen. The high-resolution data set used for refinement was collected at beamline 711 at the MAX Laboratory (Lund, Sweden) synchrotron radiation source. The crystal was transferred to mother liquor containing 20% ethylene glycol before freezing. All data were processed with DENZO and SCALEPACK (24Otwinowski Z. Data Collection and Processing. SERC Daresbury Laboratory, Warrington, United Kingdom1993: 56-62Google Scholar). The structure was solved by multiple isomorphous replacement based on five heavy metal derivatives (Table I). Location of metal binding sites and phase calculations were performed using SOLVE (25Terwilliger T.C. Berendzen J. Acta Crystallogr. Sect. D Biol. Crystallogr. 1999; 55: 849-861Crossref PubMed Scopus (3220) Google Scholar) with a native data set collected at the home source as a reference. The initial electron density map calculated at 2.7 Å was of sufficient quality to allow automated model building of most residues in the structure and phase extension to 1.46 Å using wARP (26Perakis A. Morris R. Lamzin V.S. Nat. Struct. Biol. 1999; 6: 458-464Crossref PubMed Scopus (2564) Google Scholar). This procedure was followed by multiple cycles of refinement in crystallography NMR software (CNS) (27Brünger A.T. Adams P.D. Clore G.M. DeLano W.L. Gros P. Grosse-Kunstleve R.W. Jiang J.-S. Kuszewski J. Nilges M. Pannu N.S. Read R.J. Rice L.M. Simonson T. Warren G.L. Acta Crystallogr. Sect. D Biol. Crystallogr. 1999; 54: 905-921Crossref Scopus (16967) Google Scholar) using a maximum likelihood target and bulk solvent correction and using model building in O (28Jones T.A. Zou J.-Y. Cowman S.W. Kjeldgaard M. Acta Crystallogr. Sect. A Found. Crystallogr. 1991; 47: 110-119Crossref PubMed Scopus (13011) Google Scholar). The final cycles of refinement were performed using Refmac5 (29Murshudov G.N. Lebedev A. Vagin A.A. Wilson K.S. Dodson E.J. Acta Crystallogr. Sect. D Biol. Crystallogr. 1999; 55: 247-255Crossref PubMed Scopus (1010) Google Scholar) because it led to faster convergence and lower R factor and Rfreevalues. Crystallographic data have been deposited at the PDB (accession code 1GV9 for the coordinate entry and accession code R1GV9SF for the structure factors).Table ISummary of the data collection, phasing, and refinement statisticsA. Data collection and phasing statisticsDatasetdminCompletenessUnique reflectionsRedundancyRsym1-aRsym = ∑n∑i|Ih,i−<Ih>|/IhIiIh,i.I/σSitesPhasing power1-bPhasing power = <Fh>/ɛ, calculated for data between 25 Å and the highest resolution of each derivative data set and given for centric/acentric reflections, respectively.Å%Native 11.46 (1.51–1.46)1-cNumbers in parentheses refer to the highest resolution shells.95.6 (84.6)44,7064.54.1 (11.9)22.2 (7.3)Native 22.18 (2.26–2.18)98.5 (85.3)14,6477.64.0 (8.9)44.5 (19.0)KAu(CN)22.89 (2.99–2.89)95.5 (97.9)56122.72.6 (4.1)35.2 (25.2)20.41 /0.40K2Pt(CN)42.98 (3.09–2.98)97.4 (77.8)57334.16.9 (13.4)17.0 (7.8)50.60 /0.77K2PtCl42.44 (2.53–2.44)91.3 (82.3)96893.55.7 (13.1)21.2 (12.0)40.77 /0.84HgCl22.89 (2.99–2.89)96.5 (89.0)61322.85.1 (8.1)18.3 (10.4)20.72 /0.79PIP2.62 (2.71–2.62)88.9 (81.0)76093.68.2 (14.9)13.4 (7.0)11.61 /1.75Figure of merit to 2.7 Å before/after solvent flattening with RESOLVE0.54 /0.71B. Refinement statisticsResolution: 25–1.46 ÅModel geometryProtein atoms1754Solvent atoms601Bond length r.m.s.d from ideal1-dr.m.s.d., root mean square deviation.0.010Bond angle r.m.s.d from ideal1.49No. of reflections Work set42,377Ramachandran plot1-eAs defined in PROCHECK. Test set2195Rfree0.211Percentage in most favored regions89.4Rcyst0.191Percentage in additional allowed regions10.6Percentage in generously allowed regions0.0Average B (Å) Protein15.20 Solvent27.08 Sulfates28.56Anisotropic temperature factor (Å)2Main chain bond-related B r.m.s.d0.804B11 = 0.84Main chain angle-related B r.m.s.d1.518B22 = −0.59Side chain bond-related B r.m.s.d1.890B33 = −0.25Side chain angle-related B r.m.s.d3.0861-a Rsym = ∑n∑i|Ih,i−<Ih>|/IhIiIh,i.1-b Phasing power = <Fh>/ɛ, calculated for data between 25 Å and the highest resolution of each derivative data set and given for centric/acentric reflections, respectively.1-c Numbers in parentheses refer to the highest resolution shells.1-d r.m.s.d., root mean square deviation.1-e As defined in PROCHECK. Open table in a new tab The crystal structure of the CRD of p58 was solved by multiple isomorphous replacement (TableI). The initial electron density map was of sufficient quality to allow tracing of most residues in the structure. The model was subsequently refined to Rwork/Rfree values of 19.1 and 21.1%, respectively. All data collection, phasing, and refinement statistics are summarized in Table I. The final model contains residues 50–277 of p58, 197 water molecules and 2 sulfate ions. The region corresponding to residues 1–30 constitutes the signal sequence, which is cleaved upon secretion of the protein into the medium and is therefore not present in the mature form of the protein. Residues 31–49 and the 6xHis tag inserted between residues 34 and 35 (23) are not visible in the electron density maps. There is also no density for the final 8 residues of the construct and for residues 165–169 of the p58 sequence, which presumably are part of a flexible loop. The CRD domain of p58 has an overall globular shape and is composed of 15 β-strands, a small α helix, and one turn of 310 helix (Fig. 1). Two major twisted antiparallel β-sheets, one seven-stranded (major) β-sheet, and one six-stranded (minor) β sheet pack against each other, forming a β-sandwich, in a variation of the jelly roll fold. The N terminus starts with two short β-strands (β1a and β1b) separated by a turn of 310 helix. This structural motif is replaced by a single and longer strand in other lectin structures; therefore, we refer to it as β1 in p58. This is the first strand of the minor β-sheet. It is followed by a long loop and the first strand (β2) of the major β-sheet. A β-hairpin (strands β3 and β4) is inserted between β2 and the second strand of the major β-sheet (β5). From here, the chain makes an excursion into the opposite (minor) β-sheet, contributing one strand (β6) before returning to the major β-sheet and forming four antiparallel strands (β7−β10). A loop followed by an α helix of two turns is inserted between β9 and β10. The chain then crosses over to the minor β-sheet, contributing three antiparallel strands (β11−β13). The last pair of strands is split between the two sheets, with β14 in the major β-sheet and β15 in the minor β-sheet. The N and C termini of the polypeptide chain are close to each other in space. The two β-sheets are curved, giving rise to a concave surface of the molecule on the side of the major β-sheet and a convex surface on the side of the minor β-sheet. A cleft is formed by a 15-residue-long loop between strands β7 and β8 of the major β-sheet, by the α helix and the loop preceding it. Residues Cys198 (strand β-10, major β-sheet) and Cys238 (strand β13, minor β-sheet) form a disulfide bond. Two peptide bonds are observed in cis-conformation: one between residues Ala128 and Asp129 at the entrance of strand 9, and the other between Gly62 and Pro63at the beginning of the loop joining strands β1b and β2. Based on their weak homology to plant lectins, it has been proposed (14Fiedler K. Simons K. Cell. 1994; 77: 625-626Abstract Full Text PDF PubMed Scopus (133) Google Scholar) that ERGIC-53 and VIP-36 define a new class of animal lectins in the secretory pathway. It is thought that these proteins function as sorting receptors for glycoproteins exiting the ER (22Hauri H.-P. Kappeler F. Andersson H. Appenzeller C. J. Cell Sci. 2000; 113: 587-596Crossref PubMed Google Scholar). Orthologues of this gene have been found in several organisms. Moreover, two other genes named ERGIC-53-like (ERGL) and GP36b have recently been identified as displaying significant homology to p58/ERGIC-53 (30Yerushalmi N. Keppler-Hafkemeyer A. Vasmatzis G. Liu X.F. Olsson P. Bera T.K. Duray P. Lee B. Pastan I. Gene (Amst.). 2001; 265: 55-60Crossref PubMed Scopus (27) Google Scholar), 2E. Hartmann, B. Reimann, D. Goerlich, T. A. Rapoport, and S. Prehn, unpublished results, GenBankTMaccession number U10362.suggesting that their products might also act as cargo receptors for glycoproteins. The fold observed here is most likely conserved in all ERGIC-53-like proteins recognized thus far because they share significant sequence identity (25–35%) and align very well in the region spanning the CRD (Fig. 2). Comparisons of the structure described here against the PDB data base using the DALI server (32Holm L. Sander C. J. Mol. Biol. 1993; 233: 123-138Crossref PubMed Scopus (3565) Google Scholar) and the program TOP (33Lu G. J. Appl. Crystallogr. 2000; 33: 176-183Crossref Scopus (193) Google Scholar) revealed that the CRD of p58 is structurally most similar to the leguminous lectins (TableII). Most leguminous lectins have a core structure composed of a β-sandwich with a concave face comprising seven β-strands and a convex face comprising six β-strands (Table II). Despite the fact that the sequence identities between p58/ERGIC-53 and leguminous lectins are generally <20%, the core of these structures shares the same basic architecture, and the secondary structure elements of p58 superimpose quite well with those of the leguminous lectins. The sugar binding sites in these lectins are all located on the concave β-sheet, with most of the residues that participate in ligand binding coming from the loops between strands at the top of the sheet.Table IIStructural homologues to p582-aThe structures were compared to that of p58 using the TOP program; the default parameters were used.ProteinCα2-bNumber of Cα atoms superimposed between the two structures.Rmsd2-cRmsd, root mean square deviation of superimposed Cα atoms.Identity2-dThe sequence identity is given between the number of Cα atoms superimposed.Concave β-sheetConvex β-sheetPDB code (ref. no.)Å%Pea lectin1461.3218.5761RIN(37Bourne Y. Roussel A. Frey M. Rouge P. Fontecilla-Camps J.C. Cambillau C. Proteins. 1990; 8: 365-376Crossref PubMed Scopus (118) Google Scholar)Lentil lectin1431.2918.9761LES (39Casset F. Hamelryck T. Loris R. Brisson J.R. Tellier C. Dao-Thi M.H. Wyns L. Poortmans F. Perez S. Imberty A. J. Biol. Chem. 1995; 270: 25619-25628Abstract Full Text Full Text PDF PubMed Scopus (68) Google Scholar)Isolectin-11411.2517.7761LOB (38Rini J.M. Hardman K.D. Einspahr H. Suddath F.L. Carver J.P. J. Biol. Chem. 1993; 268: 10126-10132Abstract Full Text PDF PubMed Google Scholar)Concanavalin A1351.2522.2765CNA (40Naismith J.H. Emmerich C. Habash J. Harrop S.J. Helliwell J.R. Hunter W.N. Raftery J. Kalb A.J. Yariv J. Acta Crystallogr. Sect. D Biol. Crystallogr. 1994; 50: 847-858Crossref PubMed Scopus (166) Google Scholar)Lectin IV1321.2925.0761LED (41Delbaere L.T.J. Vandoselaar M. Prasas M.L. Quail J.W. Wilson K.S. Dauter Z. J. Mol. Biol. 1993; 230: 950-965Crossref PubMed Scopus (151) Google Scholar)κ-Karragenase1141.4614.9651DYP (42Michel G. Chantalat L. Duee E. Barbeyron T. Henrissat B. Kloareg B. Dideberg O. Structure. 2001; 9: 513-525Abstract Full Text Full Text PDF PubMed Scopus (170) Google Scholar)Calnexin891.936.7761JHN (34Schrag J.D. Li J.J.M. Bergeron Y. Borisova S. Hahn M. Thomas D.Y. Cygler M. Mol. Cell. 2001; 8: 633-644Abstract Full Text Full Text PDF PubMed Scopus (318) Google Scholar)Galectin-3801.5213.8651A3K (35Seetharaman J. Kanigsberg A. Slaaby R. Leffler H. Barondes S. Rini J.M. J. Biol. Chem. 1998; 273: 13047-13052Abstract Full Text Full Text PDF PubMed Scopus (370) Google Scholar)Neurexin 1β871.9010.3771C4R (36Rudenko G. Nguyen T. Chelliah Y. Sudhof T.C. Deisenhofer J. Cell. 1999; 99: 93-101Abstract Full Text Full Text PDF PubMed Scopus (116) Google Scholar)2-a The structures were compared to that of p58 using the TOP program; the default parameters were used.2-b Number of Cα atoms superimposed between the two structures.2-c Rmsd, root mean square deviation of superimposed Cα atoms.2-d The sequence identity is given between the number of Cα atoms superimposed. Open table in a new tab Similarity is also found between p58 and other animal proteins, including lectins such as calnexin and galectin-3, as well as the ligand-binding domain of neurexin 1β (Table II). Calnexin is also a calcium-dependent lectin that resides in the ER and is involved in quality control mechanisms (34Schrag J.D. Li J.J.M. Bergeron Y. Borisova S. Hahn M. Thomas D.Y. Cygler M. Mol. Cell. 2001; 8: 633-644Abstract Full Text Full Text PDF PubMed Scopus (318) Google Scholar). When compared, p58 and calnexin have a CRD where the β-sheets have similar arrangements and show helical insertions in the same region. Despite the low sequence identity between the two proteins (Table II), 2 aspartate residues, which coordinate Ca2+ in the leguminous lectin structures, are conserved and occupy similar positions in both animal lectin structures. Galectin-3 belongs to a family of calcium-independent animal lectins that are predominantly cytoplasmic (35Seetharaman J. Kanigsberg A. Slaaby R. Leffler H. Barondes S. Rini J.M. J. Biol. Chem. 1998; 273: 13047-13052Abstract Full Text Full Text PDF PubMed Scopus (370) Google Scholar). Compared with p58, the β-sheets and loops of galectin-3 are smaller, despite their similar arrangement. Neurexin 1β belongs to a family of proteins expressed in hundreds of isoforms in neuronal tissues and thought to function as cell recognition molecules. Although ligands for these molecules have still not been identified, it has been proposed that they use the same binding fold and surface as the leguminous lectins to interact with cell surface molecules (36Rudenko G. Nguyen T. Chelliah Y. Sudhof T.C. Deisenhofer J. Cell. 1999; 99: 93-101Abstract Full Text Full Text PDF PubMed Scopus (116) Google Scholar). However, p58 and neurexin 1β superimpose poorly in the region of the putative ligand binding site, and there is no similarity between residues thought to be involved in ligand binding in p58/ERGIC-53 and the corresponding residues of neurexin 1β. Alignment of p58/ERGIC-53 and related sequences from different organisms within the animal kingdom reveals a high degree of sequence identity (Fig. 2). The sequence conservation is well distributed throughout the polypeptide chain. Highly conserved residues include: (i) tryptophans (Trp51, Trp78, Trp110, and Trp128) forming a hydrophobic ladder that runs through the hydrophobic core of the protein, (ii) glycines in loops between the β-strands, (iii) the two disulfide-bonded cysteines, and (iv) the proline observed incis-conformation in p58/ERGIC-53 and conserved in all sequences of this domain family. It is therefore likely that the overall structure observed here is conserved in other p58/ERGIC-53-like domains. Two patches of conserved residues on opposite sides of the molecule are evident, indicating that conserved residues are not confined to the hydrophobic core of the protein (Fig.3).Figure 3Surface features of p58. Aand C, electrostatic surface potential of p58. The two views are related by ∼180° rotation around a vertical axis in the plane of the paper and show the concave (A) and convex (C) faces of the molecule. Negative potential is denoted byred, and positive potential is denoted by blue. An arrow indicates the position of the putative ligand binding site. The maps are contoured at the 10kT level. Band D, sequence conservation projected onto the accessible surface of p58. Colors for the sequence conservation are as described in the Fig. 2 legend. Residues Asp129 and Asn164, for which mutagenesis studies have been carried out, are shown in orange and dark blue, respectively. The orientations are as described in A andC. This figure was prepared using GRASP (31Nicholls A. Sharp K.A. Honig B. Proteins. 1991; 11: 281-296Crossref PubMed Scopus (5316) Google Scholar).View Large Image Figure ViewerDownload Hi-res image Download (PPT) The oligomeric form of p58/ERGIC-53 binds to mannose-substituted (Man1) resins, although mannose monosaccharide is probably not the ligand of p58in vivo (19Appenzeller C. Andersson H. Kappeler F. Hauri H.-P. Nat. Cell Biol. 1999; 1: 330-334Crossref PubMed Scopus (273) Google Scholar). Rather, it is thought that p58/ERGIC-53 recognizes Man9 on glycoproteins (6Pimpaneau V. Midoux P. Monsigny M. Roche A.C. Carbohydr. Res. 1991; 213: 95-108Crossref PubMed Scopus (24) Google Scholar, 22Hauri H.-P. Kappeler F. Andersson H. Appenzeller C. J. Cell Sci. 2000; 113: 587-596Crossref PubMed Google Scholar). Mutagenesis studies have implicated 2 residues, Asp129 and Asn164, to be required for binding of ERGIC-53 to mannose-substituted resins (13Itin C. Roche A.-C. Monsigny M. Hauri H.-P. Mol. Biol. Cell. 1996; 7: 483-493Crossref PubMed Scopus (154) Google Scholar). Due to the presence of EDTA in the crystallization conditions, no Ca2+ ions are observed in the putative ligand binding site of the p58 structure when compared with those of leguminous lectins. However, similarities are observed between these binding site structures (Fig.4): residues Asp129 and Asp160 in p58 are in positions similar to those of the equivalent Asp81 and Asp121 from theLathyrus ochrus and pea lectin structures in complex with mannose (PDB codes 1RIN and 1LOB, respectively; hereafter, we refer to these two proteins by their PDB codes) (37Bourne Y. Roussel A. Frey M. Rouge P. Fontecilla-Camps J.C. Cambillau C. Proteins. 1990; 8: 365-376Crossref PubMed Scopus (118) Google Scholar, 38Rini J.M. Hardman K.D. Einspahr H. Suddath F.L. Carver J.P. J. Biol. Chem. 1993; 268: 10126-10132Abstract Full Text PDF PubMed Google Scholar). Both these residues coordinate the Ca2+ ion, and Asp81 also binds mannose in these complexes. The peptide bond between residues Ala128 and Asp129 is in thecis-conformation in p58, as in the leguminous lectins. This is essential for the correct geometry of the Ca2+ binding site and for sugar binding in these lectins (37Bourne Y. Roussel A. Frey M. Rouge P. Fontecilla-Camps J.C. Cambillau C. Proteins. 1990; 8: 365-376Crossref PubMed Scopus (118) Google Scholar). Major differences observed between the binding sites from p58, 1LOB, and 1RIN include: (i) a different conformation of Asn170 in p58 as compared with the equivalent Asp129 in 1RIN and1LOB, due mostly to a disordered region between residues 165 and 169 of p58 (Fig. 1), (ii) positioning of the side chain of Asn164∼12.0 Å away from the equivalent Asn125 in 1RIN and 1LOB(Fig. 4), and (iii) substitution of residues Glu119 and His136, which are involved in Mn2+ coordination in leguminous lectins, by Phe158 and Ala172 in p58, which probably renders it unable to bind Mn2+ ions. Of the other residues of 1LOB and 1RIN that interact with mannose, Ala210 and Glu211 superimpose well on the equivalent residues Gly259 and Gly260 from p58, whereas the third residue is part of a loop and is located around 6 Å from its counterpart in 1LOB and 1RIN. Recent data indicate that ERGIC-53 acts as a cargo receptor for a subset of glycoproteins through recognition of mannose residues in their sugar moieties (19Appenzeller C. Andersson H. Kappeler F. Hauri H.-P. Nat. Cell Biol. 1999; 1: 330-334Crossref PubMed Scopus (273) Google Scholar, 22Hauri H.-P. Kappeler F. Andersson H. Appenzeller C. J. Cell Sci. 2000; 113: 587-596Crossref PubMed Google Scholar). Because this is a selective process, and all glycoproteins have identical sugar structures while still in the ER, specific recognition of a small subset of glycoproteins is most likely also dependent in part on characteristics intrinsic to the recognized proteins other than their sugar structure. Electrostatic surface charge calculations show a marked charge polarity on the surface of p58. A deep, negatively charged pocket is situated adjacent to the residues that are thought to be involved in calcium coordination and mannose binding (Fig. 3A). The presence of a Ca2+ ion would reduce the negative charge in this pocket, but electrostatic calculations suggest that it would still have a predominantly negative character (data not shown). The sequence conservation in the vicinity as well as inside of the negatively charged pocket is quite strong (Fig. 3B), suggesting that this region and its intrinsic electrostatic character are important for protein-protein interactions. On the side of p58 opposite to the ligand binding site, there is a surface patch made up of residues conserved only in p58/ERGIC-53 orthologues (Fig. 3D). It maps to the convex β-sheet in p58 and has well-balanced charge distribution (Fig. 3C). Due to its conservation only in p58/ERGIC-53 orthologues, it may be involved in protein-protein interactions that are unique to p58/ERGIC-53, such as receptor-cargo binding or contacting neighboring molecules in the formation of oligomers. Cargo proteins might use two different and opposing faces of the p58 β-sandwich for complex formation. Whereas the carbohydrate moiety interacts with the negatively charged pocket on one side of p58/ERGIC-53, another region, conferring specificity for the cargo protein, could bind to the conserved surface patch on the other side of p58/ERGIC-53. This would be reminiscent of the interactions between calnexin and its substrates using both its arm and lectin domains (34Schrag J.D. Li J.J.M. Bergeron Y. Borisova S. Hahn M. Thomas D.Y. Cygler M. Mol. Cell. 2001; 8: 633-644Abstract Full Text Full Text PDF PubMed Scopus (318) Google Scholar). p58/ERGIC-53 oligomerizes into dimers and hexamers in contrast to other members of this family, which are monomeric. Therefore, involvement of this surface patch in oligomerization would also explain its conservation only in p58/ERGIC-53 orthologues. In summary, we have demonstrated that the CRD of p58/ERGIC-53 is an example of the utilization of the plant lectin fold within the quality control system in the ER. The fold of the structure reported here is a versatile scaffold for carbohydrate-mediated ligand-receptor interactions. The structural conservation between the CRD domain of p58 and calnexin highlights the importance of quality control mechanisms for the proper maturation of proteins in the ER. It is therefore likely that this domain was present in evolution before the separation of the plant and animal kingdoms. The structure presented here, together with functional data on p58/ERGIC-53, suggests that the leguminous lectin fold is also used for Ca2+-dependent recognition of carbohydrate structures in animals, reinforcing the proposition that these proteins constitute a new class of animal calcium-dependent lectins. We acknowledge access to synchrotron radiation at the MAX Laboratory, beamline 711. We thank Tatyana Sandalova for help with data collection and refinement and Cristofer Enroth for help with wARP." @default.
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• W2077047894 title "Crystal Structure of the Carbohydrate Recognition Domain of p58/ERGIC-53, a Protein Involved in Glycoprotein Export from the Endoplasmic Reticulum" @default.
• W2077047894 cites W1499181135 @default.
• W2077047894 cites W1503628938 @default.
• W2077047894 cites W1520730837 @default.
• W2077047894 cites W1757503351 @default.
• W2077047894 cites W1963051938 @default.
• W2077047894 cites W1965277349 @default.
• W2077047894 cites W1966398753 @default.
• W2077047894 cites W1969244755 @default.
• W2077047894 cites W1975698355 @default.
• W2077047894 cites W1979110812 @default.
• W2077047894 cites W1980253910 @default.
• W2077047894 cites W1986191025 @default.
• W2077047894 cites W1990049796 @default.
• W2077047894 cites W1991736711 @default.
• W2077047894 cites W1995017064 @default.
• W2077047894 cites W1999590520 @default.
• W2077047894 cites W2002018183 @default.
• W2077047894 cites W2009388678 @default.
• W2077047894 cites W2013083986 @default.
• W2077047894 cites W2017765863 @default.
• W2077047894 cites W2017902030 @default.
• W2077047894 cites W2022058405 @default.
• W2077047894 cites W2038063690 @default.
• W2077047894 cites W2042471904 @default.
• W2077047894 cites W2045571454 @default.
• W2077047894 cites W2045893780 @default.
• W2077047894 cites W2059439524 @default.
• W2077047894 cites W2062613923 @default.
• W2077047894 cites W2073253049 @default.
• W2077047894 cites W2080528351 @default.
• W2077047894 cites W2082103548 @default.
• W2077047894 cites W2106315897 @default.
• W2077047894 cites W2106882534 @default.
• W2077047894 cites W2108149965 @default.
• W2077047894 cites W2123970552 @default.
• W2077047894 cites W2135998691 @default.
• W2077047894 cites W2142329004 @default.
• W2077047894 cites W2158141470 @default.
• W2077047894 cites W2160951461 @default.
• W2077047894 cites W2165552942 @default.
• W2077047894 cites W2167725841 @default.
• W2077047894 cites W2169825983 @default.
• W2077047894 cites W2171451736 @default.
• W2077047894 cites W4252976088 @default.
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