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PMID:12270713

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Citation

Ellgaard, L, Bettendorff, P, Braun, D, Herrmann, T, Fiorito, F, Jelesarov, I, Güntert, P, Helenius, A and Wüthrich, K (2002) NMR structures of 36 and 73-residue fragments of the calreticulin P-domain. J. Mol. Biol. 322:773-84

Abstract

Calreticulin (CRT) is an abundant, soluble molecular chaperone of the endoplasmic reticulum. Similar to its membrane-bound homolog calnexin (CNX), it is a lectin that promotes the folding of proteins carrying N-linked glycans. Both proteins cooperate with an associated co-chaperone, the thiol-disulfide oxidoreductase ERp57. This enzyme catalyzes the formation of disulfide bonds in CNX and CRT-bound glycoprotein substrates. Previously, we solved the NMR structure of the central proline-rich P-domain of CRT comprising residues 189-288. This structure shows an extended hairpin topology, with three short anti-parallel beta-sheets, three small hydrophobic clusters, and one helical turn at the tip of the hairpin. We further demonstrated that the residues 225-251 at the tip of the CRT P-domain are involved in direct contacts with ERp57. Here, we show that the CRT P-domain fragment CRT(221-256) constitutes an autonomous folding unit, and has a structure highly similar to that of the corresponding region in CRT(189-288). Of the 36 residues present in CRT(221-256), 32 form a well-structured core, making this fragment one of the smallest known natural sequences to form a stable non-helical fold in the absence of disulfide bonds or tightly bound metal ions. CRT(221-256) comprises all the residues of the intact P-domain that were shown to interact with ERp57. Isothermal titration microcalorimetry (ITC) now showed affinity of this fragment for ERp57 similar to that of the intact P-domain, demonstrating that CRT(221-256) may be used as a low molecular mass mimic of CRT for further investigations of the interaction with ERp57. We also solved the NMR structure of the 73-residue fragment CRT(189-261), in which the tip of the hairpin and the first beta-sheet are well structured, but the residues 189-213 are disordered, presumably due to lack of stabilizing interactions across the hairpin.

Links

PubMed

Keywords

Amino Acid Sequence; Animals; Calreticulin/chemistry; Calreticulin/genetics; Computer Simulation; Gene Expression; Models, Molecular; Molecular Chaperones/chemistry; Molecular Chaperones/genetics; Molecular Sequence Data; Nuclear Magnetic Resonance, Biomolecular; Peptide Fragments/chemistry; Peptide Fragments/genetics; Protein Structure, Tertiary; Rats; Solutions

Significance

Annotations

Gene product Qualifier GO Term Evidence Code with/from Aspect Extension Notes Status

RAT:CALR

located_in

GO:0005783: endoplasmic reticulum

ECO:0000304: author statement supported by traceable reference used in manual assertion

C

Seeded From UniProt

complete

RAT:CALR

part_of

GO:0005783: endoplasmic reticulum

ECO:0000304: author statement supported by traceable reference used in manual assertion

C

Seeded From UniProt

complete

RAT:CALR

involved_in

GO:0006457: protein folding

ECO:0000304: author statement supported by traceable reference used in manual assertion

P

Seeded From UniProt

complete


See also

References

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