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PDBsum entry 1ttl
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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 alpha2delta auxiliary subunit reduces affinity of omega-Conotoxins for recombinant n-Type (cav2.2) calcium channels.
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Authors
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J.Mould,
T.Yasuda,
C.I.Schroeder,
A.M.Beedle,
C.J.Doering,
G.W.Zamponi,
D.J.Adams,
R.J.Lewis.
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Ref.
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J Biol Chem, 2004,
279,
34705-34714.
[DOI no: ]
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PubMed id
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Abstract
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The omega-conotoxins from fish-hunting cone snails are potent inhibitors of
voltage-gated calcium channels. The omega-conotoxins MVIIA and CVID are
selective N-type calcium channel inhibitors with potential in the treatment of
chronic pain. The beta and alpha(2)delta-1 auxiliary subunits influence the
expression and characteristics of the alpha(1B) subunit of N-type channels and
are differentially regulated in disease states, including pain. In this study,
we examined the influence of these auxiliary subunits on the ability of the
omega-conotoxins GVIA, MVIIA, CVID and analogues to inhibit peripheral and
central forms of the rat N-type channels. Although the beta3 subunit had little
influence on the on- and off-rates of omega-conotoxins, coexpression of
alpha(2)delta with alpha(1B) significantly reduced on-rates and equilibrium
inhibition at both the central and peripheral isoforms of the N-type channels.
The alpha(2)delta also enhanced the selectivity of MVIIA, but not CVID, for the
central isoform. Similar but less pronounced trends were also observed for
N-type channels expressed in human embryonic kidney cells. The influence of
alpha(2)delta was not affected by oocyte deglycosylation. The extent of recovery
from the omega-conotoxin block was least for GVIA, intermediate for MVIIA, and
almost complete for CVID. Application of a hyperpolarizing holding potential
(-120 mV) did not significantly enhance the extent of CVID recovery.
Interestingly, [R10K]MVIIA and [O10K]GVIA had greater recovery from the block,
whereas [K10R]CVID had reduced recovery from the block, indicating that position
10 had an important influence on the extent of omega-conotoxin reversibility.
Recovery from CVID block was reduced in the presence of alpha(2)delta in human
embryonic kidney cells and in oocytes expressing alpha(1B-b). These results may
have implications for the antinociceptive properties of omega-conotoxins, given
that the alpha(2)delta subunit is up-regulated in certain pain states.
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Figure 4.
FIG. 4. Potency of  -conotoxins at  [1B]
expressed in the presence or absence of  3 and [2]  . A,
potency estimates (K[d]) obtained from kinetic data in Tables I
and II. B, potency estimates obtained from
concentration-response curves at [1B-d] (n = 4-8 using
six batches of oocytes) produced similar results. Current
inhibition for [1B-d] + 3 or [1B-d] +
3 +
[2]
was
evaluated at 7 or 10 min after starting toxin perfusion,
respectively (see Fig. 5). Data indicated by the diamonds are
from two batches of oocytes showing an unusually high
sensitivity of [1B-d] + 3 to the
block by -conotoxin MVIIA (n =
4-7). Error bars show S.E.
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Figure 8.
FIG. 8. 1H NMR studies. A, H chemical shift for the
three residue 10-substituted -conotoxins and their
native counterparts. B, three-dimensional solution structures of
MVIIA (accession number RCSB022889; Protein Data Bank (PDB) code
1TTK [PDB]
) (15), [R10K]MVIIA (accession number RCSB02280; PDB code 1TT3
[PDB]
) (15), GVIA (accession number RCSB0228890; PDB code 1TTL [PDB]
), and [O10K]GVIA (accession number RCSB022862; PDB code 1TR6
[PDB]
). Superimposition of the 20 lowest energy conformations across
the entire backbone are shown with the exception of [O10K]GVIA,
which was superimposed over residues 1-9 and 15-27.
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The above figures are
reprinted
by permission from the ASBMB:
J Biol Chem
(2004,
279,
34705-34714)
copyright 2004.
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