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PDBsum entry 5aht
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References listed in PDB file
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Key reference
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Title
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The nedd4-1 ww domain recognizes the py motif peptide through coupled folding and binding equilibria.
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Authors
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V.Panwalkar,
P.Neudecker,
M.Schmitz,
J.Lecher,
M.Schulte,
K.Medini,
M.Stoldt,
M.A.Brimble,
D.Willbold,
A.J.Dingley.
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Ref.
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Biochemistry, 2016,
55,
659-674.
[DOI no: ]
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PubMed id
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Abstract
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The four WW domains of human Nedd4-1 (neuronal precursor cell expressed
developmentally downregulated gene 4-1) interact with the PPxY (PY) motifs of
the human epithelial Na(+) channel (hENaC) subunits, with the third WW domain
(WW3*) showing the highest affinity. We have shown previously that the α-hENaC
PY motif binding interface of WW3* undergoes conformational exchange on the
millisecond time scale, indicating that conformational sampling plays a role in
peptide recognition. To further understand this role, the structure and dynamics
of hNedd4-1 WW3* were investigated. The nuclear Overhauser effect-derived
structure of apo-WW3* resembles the domain in complex with the α-hENaC peptide,
although particular side chain conformations change upon peptide binding, which
was further investigated by molecular dynamics simulations. Model-free analysis
of the (15)N nuclear magnetic resonance spin relaxation data showed that the apo
and peptide-bound states of WW3* have similar backbone picosecond to nanosecond
time scale dynamics. However, apo-WW3* exhibits pronounced chemical exchange on
the millisecond time scale that is quenched upon peptide binding. (1)HN and
(15)N Carr-Purcell-Meiboom-Gill (CPMG) relaxation dispersion experiments at
various temperatures revealed that apo-WW3* exists in an equilibrium between the
natively folded peptide binding-competent state and a random coil-like denatured
state. The thermodynamics of the folding equilibrium was determined by fitting a
thermal denaturation profile monitored by circular dichroism spectroscopy in
combination with the CPMG data, leading to the conclusion that the unfolded
state is populated to ∼20% at 37 °C. These results show that the binding of
the hNedd4-1 WW3* domain to α-hENaC is coupled to the folding equilibrium.
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