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Copper binding to octarepeat peptides of the prion protein monitored by mass spectrometry
Author(s) -
Whittal Randy M.,
Ball Haydn L.,
Cohen Fred E.,
Burlingame Alma L.,
Prusiner Stanley B.,
Baldwin Michael A.
Publication year - 2000
Publication title -
protein science
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 3.353
H-Index - 175
eISSN - 1469-896X
pISSN - 0961-8368
DOI - 10.1110/ps.9.2.332
Subject(s) - chemistry , peptide , electrospray ionization , crystallography , circular dichroism , histidine , cooperativity , mass spectrometry , divalent , binding site , cooperative binding , stoichiometry , ion , dissociation constant , biophysics , biochemistry , amino acid , chromatography , biology , organic chemistry , receptor
Electrospray ionization mass spectrometry (ESI‐MS) was used to measure the binding of Cu 2+ ions to synthetic peptides corresponding to sections of the sequence of the mature prion protein (PrP). ESI‐MS demonstrates that Cu 2+ is unique among divalent metal ions in binding to PrP and defines the location of the major Cu 2+ binding site as the octarepeat region in the N‐terminal domain, containing multiple copies of the repeat ProHisGlyGlyGlyTrpGlyGln. The stoichiometries of the complexes measured directly by ESI‐MS are pH dependent: a peptide containing four octarepeats chelates two Cu 2+ ions at pH 6 but four at pH 7.4. At the higher pH, the binding of multiple Cu 2+ ions occurs with a high degree of cooperativity for peptides C‐terminally extended to incorporate a fifth histidine. Dissociation constants for each Cu 2+ ion binding to the octarepeat peptides, reported here for the first time, are mostly in the low micromolar range; for the addition of the third and fourth Cu 2+ ions to the extended peptides at pH 7.4, K D 's are <100 nm. n‐terminal acetylation of the peptides caused some reduction in the stoichiometry of binding at both ph's. cu 2+ also binds to a peptide corresponding to the extreme N‐terminus of PrP that precedes the octarepeats, arguing that this region of the sequence may also make a contribution to the Cu 2+ complexation. Although the structure of the four‐octarepeat peptide is not affected by pH changes in the absence of Cu 2+ , as judged by circular dichroism, Cu 2+ binding induces a modest change at pH 6 and a major structural perturbation at pH 7.4. It is possible that PrP functions as a Cu 2+ transporter by binding Cu 2+ ions from the extracellular medium under physiologic conditions and then releasing some or all of this metal upon exposure to acidic pH in endosomes or secondary lysosomes.

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