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References listed in PDB file
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Key reference
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Title
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Inclining the purine base binding plane in protein kinase ck2 by exchanging the flanking side-Chains generates a preference for ATP as a cosubstrate.
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Authors
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C.W.Yde,
I.Ermakova,
O.G.Issinger,
K.Niefind.
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Ref.
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J Mol Biol, 2005,
347,
399-414.
[DOI no: ]
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PubMed id
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Abstract
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Protein kinase CK2 (casein kinase 2) is a highly conserved and ubiquitously
found eukaryotic serine/threonine kinase that plays a role in various cellular
key processes like proliferation, apoptosis and circadian rhythm. One of its
prominent biochemical properties is its ability to use GTP as well as ATP as a
cosubstrate (dual-cosubstrate specificity). This feature is exceptional among
eukaryotic protein kinases, and its biological significance is unknown. We
describe here a mutant of the catalytic subunit of protein kinase CK2 (CK2alpha)
from Homo sapiens (hsCK2alpha) with a clear and CK2-atypical preference for ATP
compared to GTP. This mutant was designed on the basis of several structures of
CK2alpha from Zea mays (zmCK2alpha) in complex with various ATP-competitive
ligands. A structural overlay revealed the existence of a "purine base
binding plane" harbouring the planar moiety of the respective ligand like
the purine base of ATP and GTP. This purine base binding plane is sandwiched
between the side-chains of Ile66 (Val66 in hsCK2alpha) and Met163, and it adopts
a significantly different orientation than in prominent homologues like
cAMP-dependent protein kinase (CAPK). By exchanging these two flanking amino
acids (Val66Ala, Met163Leu) in hsCK2alpha(1-335), a C-terminally truncated
variant of hsCK2alpha, the cosubstrate specificity shifted in the expected
direction so that the mutant strongly favours ATP. A structure determination of
the mutant in complex with an ATP-analogue confirmed the predicted change of the
purine base binding plane orientation. An unexpected but in retrospect plausible
consequence of the mutagenesis was, that the helix alpha D region, which is in
the direct neighbourhood of the ATP-binding site, has adopted a conformation
that is more similar to CAPK and less favourable for binding of GTP. These
findings demonstrate that CK2alpha possesses sophisticated structural
adaptations in favour of dual-cosubstrate specificity, suggesting that this
property could be of biological significance.
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Figure 2.
Figure 2. Stereo pictures of selected sections of zmCK2a.
(a) The hypothetical benzamidine molecule (covered by blue
s[a]-weighted 2F[o] -F[c] electron density; no. 2 in Table 2)
within its protein environment (green electron density). The
corresponding room temperature structure (no. 1 in Table 2)
looks essentially identical (not shown). For comparision
equivalent parts of the zmCK2a/AMPPNP complex (no. 3 in Table 2)
are drawn in black. The hypothetical benzamidine molecule and
the adenine group of the bound AMPPNP molecule are almost
co-planar but do not overlap. Two alternative side-chain
conformations of Met163 are found in structure 2 of Table 2 but
only one of these is selected when AMPPNP is bound (black bonds;
black electron density). For comparison two further side-chain
conformations of Met163 are displayed as observed in
apo-zmCK2a^21 (brown) and in a zmCK2a complex with
4,5,6,7-tetrabromo-2-benzotriazole21 (magenta colour). All
pieces of electron density are drawn with a 1s contour level.
Some hydrogen bonds are indicated with pink broken lines. (b)
The divalent sulphur atom of Met163 (structure 3 of Table 2)
attached simultaneously to two p-systems, namely the adenine
group of AMPPNP and the terminal amide group of Asn118. The
final electron density is drawn in green with a contour level of
1s.
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Figure 4.
Figure 4. Structural characterisation of the mutant
hsCK2a^1-335-V66A/M163L. (a) Stereo picture of the AMPPNP
molecule (covered by blue s[a]-weighted 2F[o] -F[c] electron
density contoured at 1s) and a part of its protein environment
(green density). For comparision the equivalent sections of the
zmCK2a/AMPPNP structure (structure no. 3 in Table 2) are drawn
with black carbon atoms. (b) The adenine group of AMPPNP and its
flanking side-chains in hsCK2a^1-335-V66A/M163L (covered by
green electron density), in hsCK2a^1-335 (blue bonds), in zmCK2a
(structure no. 3 of Table 2; black bonds) and in CAPK (magenta
bonds). (c) Main chain atom RMS deviations after superimposition
of the structures of hsCK2a^1-335 and hsCK2a^1-335-V66A/M163L.
(d) Stereo picture to illustrate the structural variation in the
helix aD region. While zmCK2a (black trace) and hsCK2a^1-335
(blue trace) deviate strongly from CAPK (magenta trace) in this
region, hsCK2a^1-335-V66A/M163L (yellow trace) is much more
similar to it. As a consequence the space at the entrance to the
purine base binding plane is restricted and the binding of
GMPPNP (and GTP) is hampered.
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The above figures are
reprinted
by permission from Elsevier:
J Mol Biol
(2005,
347,
399-414)
copyright 2005.
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Secondary reference #1
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Title
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Crystal structure of the catalytic subunit of protein kinase ck2 from zea mays at 2.1 a resolution.
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Authors
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K.Niefind,
B.Guerra,
L.A.Pinna,
O.G.Issinger,
D.Schomburg.
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Ref.
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Embo J, 1998,
17,
2451-2462.
[DOI no: ]
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PubMed id
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Figure 7.
Figure 7 Stereo illustration of the adenine moiety in the
hydrophobic purine binding pocket surrounded by the non-polar
side chains contributing to the hydrophobic character. The net
marks the hydrophobic surface of the protein matrix including
water, with the intensity of the blue colour as an indicator of
hydrophobicity. This surface was calculated with BRAGI
(Schomburg and Reichelt, 1988). For comparison, the adenine
moiety of cAPK-bound ATP is drawn (green) after global 3D-fit of
the cAPK/ATP complex on rmCK2  .
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Figure 8.
Figure 8 Helix D
region with ATP ribose anchor in cAPK (yellow), CDK2 (green),
protein kinase CK1 (red) and rmCK2 (grey)
after global 3D-fits of the structures.
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The above figures are
reproduced from the cited reference
which is an Open Access publication published by Macmillan Publishers Ltd
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Secondary reference #2
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Title
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Expression, Purification and crystallization of the catalytic subunit of protein kinase ck2 from zea mays.
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Authors
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B.Guerra,
K.Niefind,
L.A.Pinna,
D.Schomburg,
O.G.Issinger.
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Ref.
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Acta Crystallogr D Biol Crystallogr, 1998,
54,
143-145.
[DOI no: ]
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PubMed id
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Figure 2.
Figure 2 Monoclinic crystals of recombinant maize CK2  -subunit.
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The above figure is
reproduced from the cited reference
with permission from the IUCr
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Secondary reference #3
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Title
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Gtp plus water mimic ATP in the active site of protein kinase ck2.
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Authors
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K.Niefind,
M.Pütter,
B.Guerra,
O.G.Issinger,
D.Schomburg.
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Ref.
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Nat Struct Biol, 1999,
6,
1100-1103.
[DOI no: ]
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PubMed id
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Figure 2.
Figure 2. The binding site of the nucleotide purine base in
rmCK2  .
a, The AMPPNP−rmCK2 complex
and b, the GMPPNP−rmCK2 complex.
The nucleotide molecules, the two magnesium ions and some
important water molecules are shown in green electron density
and the interdomain hinge region of the protein is shown in blue
electron density. All pieces of electron density are drawn from
(2F[o] - F[c]) density maps above a level of 1.2  .
Important hydrogen bonds are marked by magenta dotted lines with
donor-acceptor distances given in Å. For comparison, the
black nucleotide is GMPPNP in (a) and AMPPNP in (b). The
comparison shown is after the three-dimensional fit of the
protein matrices together with the most important water
molecules (large black balls) in its environment.
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Figure 4.
Figure 4. Stereoview of ATP in complex with the interdomain
hinge region of CAPK. AMPPNP and GMPPNP as bound to rmCK2
are
drawn in black (AMPPNP) and gray (GMPPNP) after
three-dimensional fitting of the corresponding protein matrices
to that of CAPK. Mn^2+ (green) and Mg^2+ (black in complex with
AMPPNP, gray in complex with GMPPNP) are drawn as balls. Some
important hydrogen bonds are marked as magenta dotted lines with
distances given in Å.
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The above figures are
reproduced from the cited reference
with permission from Macmillan Publishers Ltd
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Secondary reference #4
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Title
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Crystallization and preliminary characterization of crystals of human protein kinase ck2.
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Authors
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K.Niefind,
B.Guerra,
I.Ermakowa,
O.G.Issinger.
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Ref.
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Acta Crystallogr D Biol Crystallogr, 2000,
56,
1680-1684.
[DOI no: ]
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PubMed id
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Figure 1.
Figure 1 Gel-filtration chromatograms of CK2 holoenzyme. (a)
Chimeric CK2 composed of maize CK2 and
human CK2  ;
(b) rhCK2 after partial degradation of CK2 .
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Figure 6.
Figure 6 180° self-rotation function calculated with GLRF (Tong
& Rossmann, 1997[Tong, L. & Rossmann, M. G. (1997). Methods
Enzymol. 276, 594-611.]) using reflections in the resolution
range 20-3.5 Å.
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The above figures are
reproduced from the cited reference
with permission from the IUCr
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Secondary reference #5
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Title
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Crystal structure of human protein kinase ck2: insights into basic properties of the ck2 holoenzyme.
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Authors
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K.Niefind,
B.Guerra,
I.Ermakowa,
O.G.Issinger.
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Ref.
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EMBO J, 2001,
20,
5320-5331.
[DOI no: ]
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PubMed id
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Figure 3.
Figure 3 Various aspects of the rhCK2^  structure.
(A and B) Overall shape of rhCK2^ in
a view perpendicular to the local C2 axis (A) and along this
axis (B). The two rhCK2  chains
are drawn in blue and red, the two rhCK2 subunits
in yellow and grey. (C) Structural overview of rhCK2 chain
A1 with bound AMPPNP and interdomain flexibility. The hinge axis
and the bending residues of the domain closure motion as
detected by DYNDOM (CCP4, 1994) are included. To illustrate the
interdomain flexibility, the N-terminal domains of rmCK2 (PDB
code: 1DAW) and of rhCK2 chain
A2 are shown in yellow and black, respectively, after
three-dimensional alignment of the corresponding C-terminal
domains. (D) Structural overview of rhCK2 .
The human CK2 peptide
bound to rmCK2 (black)
was taken from PDB file: 1DS5 (Battistutta et al., 2000) after
superimposition of the corresponding CK2 subunits.
(E) Intersubunit flexibility at the /
contact.
Subunit A1 is drawn with yellow colour for the C-terminal domain
and grey for the N-terminal domain. Subunit B1 bound to A1 by an
/
contact
is sketched in red. Subunit B2 is shown in blue after a
three-dimensional fit of the N-terminal domain of subunit A2
(not drawn) on that of A1.
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Figure 4.
Figure 4 Electrostatic surface of rhCK2^ .
The surface is coloured according to the electrostatic potential
ranging from deep blue (positive charge) to red (negative
charge). Atomic charges were assigned by GRASP (Nicholls et al.,
1991) using default values.
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The above figures are
reproduced from the cited reference
which is an Open Access publication published by Macmillan Publishers Ltd
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Secondary reference #6
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Title
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Expression and characterization of a recombinant maize ck-2 alpha subunit.
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Authors
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B.Boldyreff,
F.Meggio,
G.Dobrowolska,
L.A.Pinna,
O.G.Issinger.
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Ref.
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Biochim Biophys Acta, 1993,
1173,
32-38.
[DOI no: ]
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PubMed id
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Secondary reference #7
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Title
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Cloning and sequencing of the casein kinase 2 alpha subunit from zea mays.
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Authors
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G.Dobrowolska,
B.Boldyreff,
O.G.Issinger.
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Ref.
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Biochim Biophys Acta, 1991,
1129,
139-140.
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PubMed id
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