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PDBsum entry 3gin
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Metal binding protein
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PDB id
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3gin
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References listed in PDB file
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Key reference
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Title
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Structure and functional analysis of a ca2+ sensor mutant of the na+/ca2+ exchanger.
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Authors
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V.Chaptal,
M.Ottolia,
G.Mercado-Besserer,
D.A.Nicoll,
K.D.Philipson,
J.Abramson.
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Ref.
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J Biol Chem, 2009,
284,
14688-14692.
[DOI no: ]
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PubMed id
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Abstract
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The mammalian Na(+)/Ca(2+) exchanger, NCX1.1, serves as the main mechanism for
Ca(2+) efflux across the sarcolemma following cardiac contraction. In addition
to transporting Ca(2+), NCX1.1 activity is also strongly regulated by Ca(2+)
binding to two intracellular regulatory domains, CBD1 and CBD2. The structures
of both of these domains have been solved by NMR spectroscopy and x-ray
crystallography, greatly enhancing our understanding of Ca(2+) regulation.
Nevertheless, the mechanisms by which Ca(2+) regulates the exchanger remain
incompletely understood. The initial NMR study showed that the first regulatory
domain, CBD1, unfolds in the absence of regulatory Ca(2+). It was further
demonstrated that a mutation of an acidic residue involved in Ca(2+) binding,
E454K, prevents this structural unfolding. A contradictory result was recently
obtained in a second NMR study in which Ca(2+) removal merely triggered local
rearrangements of CBD1. To address this issue, we solved the crystal structure
of the E454K-CBD1 mutant and performed electrophysiological analyses of the
full-length exchanger with mutations at position 454. We show that the lysine
substitution replaces the Ca(2+) ion at position 1 of the CBD1 Ca(2+) binding
site and participates in a charge compensation mechanism. Electrophysiological
analyses show that mutations of residue Glu-454 have no impact on Ca(2+)
regulation of NCX1.1. Together, structural and mutational analyses indicate that
only two of the four Ca(2+) ions that bind to CBD1 are important for regulating
exchanger activity.
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Figure 1.
Structural comparison between WT-CBD1 (gray, PDB code: 2DPK)
and E454K-CBD1 (blue). The three Ca^2+ ions bound to E454K-CBD1
are displayed as green spheres and numbered according to WT-CBD1
numbering (5). For clarity, the β strands are labeled A–G.
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Figure 2.
Overlay of the Ca^2+ binding sites of WT-CBD1 and E454K-CBD1.
E454K-CBD1 is shown as a blue drawing, and Ca^2+ ions are
depicted as green spheres. Ca^2+-coordinating residues are
displayed as sticks colored by atom type (Cα is yellow).
Relevant metal ion coordinations and charge pairs of E454K-CBD1
are shown as black dotted lines. WT-CBD1 is shown as a gray
drawing, and equivalent residues are displayed as sticks colored
by atom type. Ca^2+ ions are numbered according to Ref. 5.
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The above figures are
reprinted
by permission from the ASBMB:
J Biol Chem
(2009,
284,
14688-14692)
copyright 2009.
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