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Molecular Origin of pH‐Dependent Fibril Formation of a Functional Amyloid
Author(s) -
McGlinchey Ryan P.,
Jiang Zhiping,
Lee Jennifer C.
Publication year - 2014
Publication title -
chembiochem
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.05
H-Index - 126
eISSN - 1439-7633
pISSN - 1439-4227
DOI - 10.1002/cbic.201402074
Subject(s) - chemistry , fibril , circular dichroism , biophysics , amyloid (mycology) , protonation , fluorescence , residue (chemistry) , mutant , protein aggregation , kinetics , fibrillogenesis , stereochemistry , crystallography , biochemistry , organic chemistry , inorganic chemistry , biology , ion , physics , quantum mechanics , gene
Fibrils derived from Pmel17 are functional amyloids upon which melanin is deposited. Fibrils of the repeat domain (RPT) of Pmel17 form under strict melanosomal pH (4.5–5.5) and completely dissolve at pH≥6. To determine which Glu residue is responsible for this reversibility, aggregation of single, double, and quadruple Ala and Gln mutants were examined by intrinsic Trp fluorescence, circular dichroism spectroscopy, and transmission electron microscopy. Charge neutralization of E404, E422, E425, or E430, which are located in the putative amyloid‐forming region, modulated aggregation kinetics. Remarkably, the removal of a single negative charge at E422, one of 16 carboxylic acids, shifted the pH dependence by a full pH unit. Mutation at E404, E425, or E430 had little to no effect. We suggest that protonation at E422 is essential for initiating amyloid formation and that the other Glu residues play an allosteric role in fibril stability.