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PDBsum entry 2noo
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References listed in PDB file
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Key reference
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Title
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Nickel binding to nika: an additional binding site reconciles spectroscopy, Calorimetry and crystallography.
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Authors
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C.Addy,
M.Ohara,
F.Kawai,
A.Kidera,
M.Ikeguchi,
S.Fuchigami,
M.Osawa,
I.Shimada,
S.Y.Park,
J.R.Tame,
J.G.Heddle.
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Ref.
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Acta Crystallogr D Biol Crystallogr, 2007,
63,
221-229.
[DOI no: ]
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PubMed id
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Abstract
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Intracellular nickel is required by Escherichia coli as a cofactor for a number
of enzymes and is necessary for anaerobic respiration. However, high
concentrations of nickel are toxic, so both import and export systems have
evolved to control the cellular level of the metal. The nik operon in E. coli
encodes a nickel-uptake system that includes the periplasmic nickel-binding
protein NikA. The crystal structures of wild-type NikA both bound to nickel and
in the apo form have been solved previously. The liganded structure appeared to
show an unusual interaction between the nickel and the protein in which no
direct bonds are formed. The highly unusual nickel coordination suggested by the
crystal structure contrasted strongly with earlier X-ray spectroscopic studies.
The known nickel-binding site has been probed by extensive mutagenesis and
isothermal titration calorimetry and it has been found that even large numbers
of disruptive mutations appear to have little effect on the nickel affinity. The
crystal structure of a binding-site mutant with nickel bound has been solved and
it is found that nickel is bound to two histidine residues at a position distant
from the previously characterized binding site. This novel site immediately
resolves the conflict between the crystal structures and other biophysical
analyses. The physiological relevance of the two binding sites is discussed.
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Figure 3.
Figure 3 ITC traces of (a) 10 µl injections of 1.1 mM
NiCl[2] into 180 µM EDTA. (b) 10 µl injections of 5
mM EDTA, 1.8 mM NiCl[2] into 180 µM NikA H56A. (c) 10
µl injections of 5 mM EDTA, 1.8 mM NiCl[2] into 180
µM wild-type NikA. (d) 10 µl injections of 5 mM
EDTA, 1.8 mM NiCl[2] into 180 µM 6-mutant NikA. 1 cal =
4.186 J.
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Figure 4.
Figure 4 (a) Crystal structure of 6-mutant NikA. The mutated
residues around the binding cleft are shown as cyan sticks. The
nickel-binding histidine residues are shown as yellow sticks.
(b) Ribbon representations of NikA (red) and 6-mutant NikA
(yellow) after alignment of the C^  backbones
of lobe 1 (residues 4-245 and 471-499) of the two proteins.
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The above figures are
reprinted
by permission from the IUCr:
Acta Crystallogr D Biol Crystallogr
(2007,
63,
221-229)
copyright 2007.
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