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Title
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Molecular basis for co-operativity in Ca2+ binding to calbindin D9k. 1H nuclear magnetic resonance studies of (Cd2+)1-bovine calbindin D9k.
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Authors
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M.Akke,
S.Forsén,
W.J.Chazin.
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Ref.
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J Mol Biol, 1991,
220,
173-189.
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PubMed id
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Abstract
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The molecular basis for the co-operativity in binding of calcium ions by bovine
calbindin D9k has been addressed by carrying out a comparative analysis of the
solution conformation and dynamics of the apo, half saturated and fully
saturated species using two-dimensional 1H nuclear magnetic resonance
spectroscopy. Since the half saturated calcium form of the protein is not
significantly populated under equilibrium conditions due to the co-operativity
in binding of calcium ions, the half saturated cadmium form of the protein has
been substituted for the calcium form. To verify that cadmium forms of calbindin
D9k represent viable models for the calcium-bound species, the fully saturated
cadmium form has been prepared and compared to the calcium-saturated protein.
Virtually complete 1H resonance assignments have been obtained for both the
(Cd2+)1 and the (Cd2+)2 states. Secondary structure elements and the global
folding pattern were determined from nuclear Overhauser effects, backbone
spin-spin coupling constants and slowly exchanging amide protons. Comparisons of
the half saturated protein with the apo and calcium-saturated forms of calbindin
D9k show that all three structures are highly similar. However, a change in the
structural and dynamic properties of the protein does occur upon binding of the
first ion; the half saturated form is found to be more similar to the
calcium-saturated form than to the apo form. These results have important
implications concerning the molecular basis for the co-operativity, and suggest
that entropic effects associated with the protein dynamics play an important
role.
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