Zn 2+ -binding in the glutamate-rich region of the intrinsically disordered protein prothymosin-alpha
JOURNAL OF BIOLOGICAL INORGANIC CHEMISTRY(2018)
Abstract
Prothymosin-α is a small, multifunctional intrinsically disordered protein associated with cell survival and proliferation which binds multiple Zn 2+ ions and undergoes partial folding. The interaction between prothymosin-α and at least two of its protein targets is significantly enhanced in the presence of Zn 2+ ions, suggesting that Zn 2+ binding plays a role in the protein’s function. The primary sequence of prothymosin-α is highly acidic, with almost 50% comprised of Asp and Glu, and is unusual for a Zn 2+ -binding protein as it lacks Cys and His residues. To gain a better understanding of the nature of the Zn 2+ -prothymosin-α interactions and the protein’s ability to discriminate Zn 2+ over other divalent cations (e.g., Ca 2+ , Co 2+ , Mg 2+ ) we synthesized a set of three model peptides and characterized the effect of metal binding using electrospray ionization mass spectrometry (ESI MS) and circular dichroism (CD) spectroscopy. ESI MS data reveal that the native peptide model of the glutamic acid rich region binds 4 Zn 2+ ions with apparent, stepwise K d values that are, at highest, in the tens of micromolar range. A peptide model with the same amino acid composition as the native sequence, but with the residues arranged randomly, showed no evidence of structural change by CD upon introduction of Zn 2+ . These results suggest that the high net negative charge of the glutamic acid-rich region of prothymosin-α is not a sufficient criterion for Zn 2+ to induce a structural change; rather, Zn 2+ binding to prothymosin-α is sequence specific, providing important insight into the behavior of intrinsically disordered proteins.
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Key words
Prothymosin-alpha,Zinc binding,Intrinsically disordered protein,Circular dichroism,Mass spectrometry
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