From the John Curtin School of Medical Research, Australian National University, P. O. Box 334, Canberra, Australian Capital Territory 2601, Australia
Received for publication, August 29, 2000, and in revised form, September 29, 2000
![]() |
ABSTRACT |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
The ubiquitous glutathione transferases (GSTs)
catalyze glutathione conjugation to many compounds and have other
diverse functions that continue to be discovered. We noticed sequence
similarities between Omega class GSTs and a nuclear chloride channel,
NCC27 (CLIC1), and show here that NCC27 belongs to the GST structural family. The structural homology prompted us to investigate whether the
human Omega class glutathione transferase GSTO1-1 forms or modulates
ion channels. We find that GSTO1-1 modulates ryanodine receptors
(RyR), which are calcium channels in the endoplasmic reticulum of
various cells. Cardiac RyR2 activity was inhibited by GSTO1-1, whereas
skeletal muscle RyR1 activity was potentiated. An enzymatically active
conformation of GSTO1-1 was required for inhibition of RyR2, and
mutation of the active site cysteine (Cys-32 Glutathione transferases
(GSTs)1 are a family of
ubiquitous intracellular enzymes that catalyze the conjugation of
glutathione to many exogenous and endogenous compounds (1). GSTs are
known to have other functions including the binding of bilirubin and carcinogens (2), the isomerization of maleylacetoacetate (3), and the
regulation of stress kinases (4), with presumably further roles yet to
be discovered. New members of the GST structural family with novel
catalytic activities and functions have recently been discovered
(5-7). For example, the Omega class glutathione transferase GSTO1-1
has a typical glutathione transferase fold but little enzymatic
activity with many conventional substrates (7). Unlike other mammalian
glutathione transferases that have active site tyrosine or serine
residues (8), GSTO1-1 has a novel active site cysteine that
participates in weak thiol transferase reactions. Although the
intracellular function of the Omega class GSTs is unknown, a member of
the Omega class is over-expressed in a radiation-resistant mouse
lymphoma cell line (9).
We used BLAST searches (10) to identify additional members of
the glutathione transferase structural family and were impressed by
sequence similarities between GSTO1-1 and the chloride
intracellular channel (CLIC) family of proteins, which are thought to
form chloride channels in intracellular membranes or to be chloride
channel modulators (11, 12). We therefore compared the structure of the
CLIC proteins and GSTO1-1 in more detail and found that NCC27 (CLIC1)
belongs to the GST structural family. Because of the structural similarity, we also examined the ability of GSTO1-1 to form or modulate ion channels. We find that GSTO1-1 modulates ryanodine receptors (RyRs), which are the calcium release channels in skeletal and cardiac sarcoplasmic reticulum (SR). There is evidence that GSTO1-1 is present in skeletal and cardiac muscle (7) and is thus
colocalized with RyRs. RyRs are also located in intracellular membranes
of a variety of cells (13) and are expressed in T- and B-lymphocytes
(14, 15). GSTO1-1 inhibited cardiac RyR2 channel activity. Modulation
of ion channels is a previously undescribed function for a GST. We
propose that GSTO1-1 plays a novel role in regulating intracellular
[Ca2+], potentially protecting cells containing RyR2 from
radiation damage (9) and apoptosis induced by Ca2+
mobilization from intracellular stores (16, 17).
Sequence Alignment--
Sequences for NCC27 (11), also known as
CLIC1 and GSTO1-1 (7), were aligned with CLUSTALW (18) with
default settings. Minor manual alterations were introduced into the
alignment, taking into consideration both the model and an alignment of
other GST family
members.2
Modelling--
Modelling was performed with the HOMOLOGY
module of the Insight II package (MSI, San Diego, CA) based on the
alignment in Fig. 1. Despite the low level of sequence identity
(~15%), visual inspection of the model confirmed that the NCC27
sequence is compatible with the canonical GST fold. Further validation
of the model was derived from applying a THREADING procedure
(19) to the NCC27 sequence that scored the highest match against
GSTO1-1 when compared with the nonredundant structure data base with a
Z score of-2.78
GSTO1-1 Preparation--
Recombinant GSTO1-1 was expressed in
Escherichia coli after cloning the cDNA into the
expression vector pQE 30 (Qiagen, Clifton Hill, Victoria,
Australia). The expressed protein has a poly-His tag and was
purified by Ni-agarose chromatography (20). The purified protein was
dialyzed into 20 mM Tris-HCl, 60 mM NaCl, 5 mM dithiothreitol, pH 8.0. The C32A mutant was
created by site-directed mutagenesis and confirmed by DNA sequence
analysis. The mutant protein was purified and stored by the same
procedures as the wild type enzyme. To block protein thiols, GSTO1-1
was diluted to 2 mg/ml in 20 mM Tris-HCl, 60 mM
NaCl, pH 8.0 containing 20 mM
N-ethylmaleimide. The protein was incubated at
20 °C for 1 h and then dialyzed extensively against 20 mM Tris-HCl, 60 mM NaCl, pH 8.0 at 4 °C.
Antiserum--
Antiserum to human GSTO1-1 was prepared by
immunizing rabbits with purified recombinant GSTO1-1 in Fruend's
adjuvant. Pre-immune serum was collected before immunization for use as
a control. The polyclonal antisera obtained recognized recombinant
GSTO1-1 and GSTO1-1 derived from human tissue extracts on Western blots.
GSTO1-1 and Lipid Bilayers--
GSTO1-1 at 21-210 µg/ml was
added to the cis side of bilayers containing
phosphatidylethanolamine, phosphatidylserine, and phosphatidylcholine
(5:3:2, w/w), and the solution was stirred for 30-90 min. Solutions
for this experiment contained the following: cis/trans, 500/50 mM NaCl; cis
and trans, 10 mM TES (pH 4.0-7.0), with or
without 1 mM CaCl2.
SR Vesicles, Ryanodine Receptor Channels, and Lipid
Bilayers--
SR vesicles were prepared from rabbit or pig skeletal
muscle or sheep cardiac muscle as described previously (21, 22). Vesicles containing RyRs were added to the cis solution for
incorporation into artificial lipid bilayers (21, 22). The cytoplasmic
surface of the incorporated SR and RyRs faced the cis
solution. RyRs were identified by their ability to conduct
Cs+ ions with a single channel conductance of 250-300
picosiemens at +40 or Solutions--
Recording solutions contained the following
(cis and trans): 230 mM cesium
methanesulfonate, 20 mM CsCl, and 10 mM TES (pH 7.4 with CsOH), with 1 mM CaCl2
(trans), and 10 Single Channel Recording and Analysis--
Single channel
recording and analysis have been described (21, 22). Mean current
(I', the average of all data points in a record) was used as
a measure of channel activity, because many bilayers contained more
than one active channel. Data are presented in Figs. 3 and 4 as
relative mean current
(I't/I'c, where
I't is the mean current under test conditions and
I'c is the control mean current). Bilayer
potential was changed between Statistics--
The significance of the difference between
control and test values was tested (a) using a Student's t
test, either 1- or 2-sided and either for independent or paired data,
as appropriate, or (b) using the nonparametric "sign" test
(23). Differences were considered to be significant when
p Sequence Alignment and Structural Modelling--
Alignment of the
human GSTO1-1 sequence and the human chloride channel protein NCC27
(Fig. 1) reveals several similarities, both in length and in the number of conserved and conservatively replaced residues. The similarity is particularly striking in the
N-terminal glutathione-binding region, with conservation of a CPF motif
that includes the active site cysteine residue of GSTO1-1. There are
four amino acid positions that are common to virtually all known
glutathione transferases (7) (Fig. 1, asterisks). These
positions include a cis Pro within the GSH binding
site and an Asp, which acts as an N-terminal cap near the start of helix 6. Two Gly residues also appear to be important for structural reasons (8). Conspicuously, all four of these residues are conserved in
the CLIC proteins. Additional sites, such as Leu-34, Asp-76, Phe-83,
Leu-84, Phe-167, and Leu-168 (Fig. 1, number signs) show a
pattern of hydrophobicity that extends throughout other GST classes.
Based on the alignment, a tentative three-dimensional structure was
predicted by homology modelling (Fig. 2).
The model is plausible because there are no uncomplexed buried charges, and most hydrophobic residues are shielded from solvent. The side of
helix 3 that forms an interface with helix 4 is quite polar, suggesting
that there may be a relative shift between the N- and C-terminal
domains with respect to other GST structures (Fig. 2). Both helix 4 and
helix 5 are likely to be kinked, with regions of irregular hydrogen
bonding. Therefore some differences between the CLIC protein and Omega
structures are predicted (7). Modelling GSH into the putative active
site reveals that a covalent bond could be formed between the GSH Cys
and Cys-24 of the CLIC protein, analogous to that observed in the Omega
class GST structure (7). Furthermore, Lys-49, Arg-51, and Arg-29 are
able to complex the negative GSH charges, whereas Glu-63 and Asp-76
complex the positive charge on the Ion Channel Modulation by GSTO1-1--
The predicted similarities
in the structure of the Omega class GSTs and the CLIC proteins led us
to examine the possibility that GSTO1-1 may behave like CLICs and act
as an ion channel and/or ion channel modulator. Although GSTO1-1 alone
did not form ion channels in lipid bilayers under the conditions we
used, we found that it modulated RyR channels. GSTO1-1 (21 µg/ml),
added to the cis (cytoplasmic)
solution, reduced cardiac RyR2 activity by about 50% in 28 of 29 bilayers (Fig. 3, b and c, and Fig.
4, b and c). A
10-fold higher GST concentration (210 µg/ml) caused similar depression of cardiac RyR activity in seven bilayers (Fig.
4b). The Ki for inhibition of RyR2
activity in four bilayers that were exposed to a series of GSTO1-1
concentrations was 1 µg/ml (Fig. 4f). The apparent
increase in activity as [GSTO1-1] increased from 21 to 210 µg/ml in Fig. 4b was not significant and may have arisen
from inclusion of data from different bilayers for the different
concentrations. The increase is not seen in the limited data set
of Fig. 4f, containing bilayers exposed to each of
the three GSTO1-1 concentrations. Control experiments showed that
buffer lacking GSTO1-1 did not affect RyR activity (Figs.
3a and 4a). Depression of RyR activity by the GST
was independent of cytoplasmic (cis) [Ca2+]
between 10
Covalent modification (oxidation) of the RyR complex by GSTO1-1 was
not responsible for the inhibition, because channel activity recovered
and was even potentiated above control levels (n = 10) when GSTO1-1 was washed out (Fig. 3b). Normal channel
activity could also be restored by addition of polyclonal anti-GSTO1-1 antiserum (n = 7), with potentiation of activity above
control levels in each case (Fig. 3c and Fig. 4,
b and c). These changes in channel activity were
a specific effect of the antibody on GSTO1-1 because (a) recovery was
not seen when 100 µl of pre-immune serum lacking antibody was added
to channels whose activity was depressed by GSTO1-1 (n = 4), and (b) 100 µl of antiserum alone did not affect RyR2 channel
activity in the absence of GSTO1-1 (Fig. 4a).
Two other kinds of experiment indicate that inhibition of RyR2 activity
by GSTO1-1 is related to its thiol transferase activity. First, when
GSTO1-1 was treated with N-ethylmaleimide (GSTO1-1-M), it
lost both its enzyme activity and its ability to inhibit RyR2 activity
(n = 8) (Figs. 3d and 4d). Indeed
channel activity was increased by GSTO1-1-M (Fig.
4d, n = 7). Control-incubated enzyme (GSTO1-1-C; see "Experimental Procedures") was then
applied, and channel activity fell to control levels (Fig.
4d). The depression may have been less than expected because
inactive GSTO1-1-M was occupying some of the inhibition sites and
preventing GSTO1-1-C binding. In the second type of experiment,
GSTO1-1 with a Cys-32 Early studies of the glutathione transferase family relied on
functional similarities to identify different family members. In
previous studies we have identified additional members of the GST
structural family by sequence alignment-based searches of the
expressed sequence tag and GenBankTM data bases (5,
7). The recently described Zeta and Omega class GSTs were identified by
this approach, and although they are clearly members of the GST
structural family, they are functionally distinct from previously
described GST classes. In the present study we utilized the Omega class
GST sequence to search for additional members of the GST structural
family. This search revealed significant sequence similarity with
members of the CLIC group of intracellular channel proteins. Detailed
examination of an alignment of the GSTO1 and NCC27 (CLIC1) sequences
revealed the conservation of several key residues that are conserved
throughout the GST structural family. Subsequent homology modelling
strongly supported the prediction that NCC27 adopts the canonical GST
fold and can be classified as a member of the GST structural family.
The formation and modulation of ion channels are novel functions never
previously associated with members of the GST family and clearly extend
the potential roles of this superfamily.
The observation that CLIC proteins probably adopt a GST fold and share
a similar N-terminal extension and `active site` motif with GSTO1
suggested that we should investigate the capacity of an Omega class GST
to form or modulate ion channels. GSTO1-1 did not form channels under
the conditions we used. However, because GSTO1-1 is strongly expressed
in both skeletal and cardiac muscle (7), we examined its capacity to
modulate RyRs. Our results clearly indicated that GSTO1-1 is able to
either inhibit or potentiate RyR calcium channels. The inhibition we
observed (a) does not depend on an oxidation reaction, (b) requires an
active enzyme conformation, and (c) is seen only in RyR2. Activation of
RyRs is independent of enzyme activity and is seen in both RyR1 and RyR2. The results suggest that RyR2 has two binding sites for GSTO1-1
and that enzyme binding to one site activates the channel, whereas
enzyme binding to the second site inhibits the channel. The RyR has a
very large cytoplasmic domain with multiple ligand binding sites, and
there are several examples of ligands with dual effects on the channel
because ligands can bind to more than one site. Examples are
Ca2+ (21), ATP (24), and peptides that correspond to parts
of the II-III loop of the dihydropyridine receptor (25).
Ion channel modulation is a novel and unrecognized role for a
GST; this is the first report of modulation of RyRs by a GST protein. RyR channels are regulated by many factors including oxidation
and reduction reactions (26). GSTO1-1 provides an additional mechanism
by which redox state might be conveyed to the channels. Cytoplasmic
[Ca2+] increases during oxidative stress, partly through
oxidative activation of RyRs (26). An increase in [GSTO1-1] during
oxidative stress could protect RyR2 from oxidation-induced activation
in all cells expressing this RyR isoform. Because CLIC proteins also form or modulate ion channels (11, 12) and have a structure similar to
that of GSTs, it may be that ion channel modulation is a common
property of this superfamily. It may be significant that Jun
N-terminal kinase signaling is regulated by GST Pi binding (4, 27) and
that the mitogen-activated protein kinase ERK7 binds to CLIC 3 (12), an
intracellular chloride channel protein shown here to be a putative
member of the GST structural family by its strong sequence similarity
to NCC27/CLIC 1. Because cardiac RyR2s form macromolecular complexes
that include protein kinase A (28), the possibility that the modulating
effects of GSTO1-1 may be mediated by interactions with the
RyR2-associated protein kinase deserves further investigation. GST
modulation of RyRs may have widespread effects. GSTO1-1 is
over-expressed in a radiation-resistant lymphoma cell line (9) and may
reduce apoptosis normally resulting from Ca2+ mobilization
from RyR-sensitive stores (17). GSTO1-1 may also protect cancer cells
from apoptosis caused by Ca2+ mobilization through RyRs
(16). Finally GSTO1-1 may modulate immune responses that depend on a
sustained increase in cytoplasmic [Ca2+] through activity
of RyRs in T- and B-lymphocytes (14, 15).
Ala) abolished
the inhibitory activity. We propose a novel role for GSTO1-1 in
protecting cells containing RyR2 from apoptosis induced by
Ca2+ mobilization from intracellular stores.
INTRODUCTION
TOP
ABSTRACT
INTRODUCTION
EXPERIMENTAL PROCEDURES
RESULTS
DISCUSSION
REFERENCES
EXPERIMENTAL PROCEDURES
TOP
ABSTRACT
INTRODUCTION
EXPERIMENTAL PROCEDURES
RESULTS
DISCUSSION
REFERENCES
40 mV and by their susceptibility to
block by 10 µM ruthenium red added to the cis
chamber at the end of each experiment.
6 or
10
5 M calcium ions
(cis) buffered using 1 mM
1,2-bis(2-aminophenoxy)-ethane-N,N,N',N'-tetraacetic acid. GSTs, buffer, and antibody were added to the cis
chamber and were "washed out" by perfusion with 10 volumes of
cis solution. Bilayer potentials are given relative to the
trans chamber. 10 and 100 µl of stock GSTO1-1 solution
(see above) were added to the cis chamber to give
final GSTO1-1 concentrations of 21 and 210 µg/ml,
respectively. Therefore 10 and 100 µl of buffer alone were
added in control experiments (see Figs. 3a and
4a).
40 and +40 mV every 30 s.
I't and I'c were measured
from 120-s recordings (2 × 30 s at
40 mV and 2 × 30 s at +40 mV) obtained under each condition, i.e.
control conditions, for each [GST] or [buffer], after antibody
addition, or after washout. Data is presented as the mean ± S.E.,
and the numbers of bilayers are given. The number of experiments
included in the average data exceeded the number of bilayers because
(a) measurements at
40 and +40 mV were included in the average, and
(b) the experiment was often repeated on one bilayer.
0.05
RESULTS
TOP
ABSTRACT
INTRODUCTION
EXPERIMENTAL PROCEDURES
RESULTS
DISCUSSION
REFERENCES
-glutamyl group of GSH.
Thus, whereas considerable caution must be exercised when interpreting
alignments with limited sequence identity, the sequence of the CLIC
pro-tein is generally consistent with the canonical GST structure,
particularly in the N-terminal domain. Additional support for
the model comes from the application of an automated
THREADING procedure to the NCC27 sequence (19). NCC27 scored the
best match with the Omega GST structure, with a Z score of
2.78 when
compared with other folds in the nonredundant structure data base.
View larger version (41K):
[in a new window]
Fig. 1.
Alignment between the CLIC protein, NCC27,
and GSTO1-1. Conserved positions between the sequences are
indicated by a vertical stroke; conservative replacements
(R-K, D-E, F-Y, S-T) are indicated by two vertical dots.
Most of these positions are also conserved within the respective CLIC
and Omega GST subfamilies. The four amino acid positions that are
highly conserved among GSTs are indicated by asterisks. All
four of these residues are conserved in the CLIC proteins. Sites
showing a pattern of hydrophobicity extending throughout other GST
classes are marked with a number sign. Putative secondary
structure segments are indicated above the alignment (h,
helix; e, strand).
View larger version (73K):
[in a new window]
Fig. 2.
A homology model of CLIC protein
NCC27 based on the alignment in Fig. 1 and the atomic
coordinates of GSTO1-1 (Protein Data Bank code
1eem). The model is low resolution but shows that the CLIC
sequence is compatible with the GSTO1-1 fold. The C-terminal extension
is modelled to form a helical structure covering the top of the active
site, a feature so far only seen in the Alpha and Omega GSTs. Placing
GSH into the putative active site of the CLIC protein reveals that it
can form interactions analogous to those observed in the Omega class
GST.
5 and 10
6
M (Fig. 3, b and c, and Fig. 4,
b and c) and independent of bilayer potential
(therefore average values in Fig. 4 include measurements at
40 and
+40 mV).
View larger version (32K):
[in a new window]
Fig. 3.
GSTO1-1 modulates RyR channels. Current
records at 40 mV are shown, as are all-points histograms of
the probability (p) of current levels, I(pA).
GSTO1-1 was added to the cis (cytoplasmic) solution.
a, a control experiment showing that RyR2 activity was
unaffected by buffer lacking GSTO1-1. In b, RyR2
activity was depressed by GSTO1-1 with 10
6
M cis Ca2+ and then recovered after
washout. In c, RyR2 activity was depressed by
GSTO1-1 with 10
5 M
cis Ca2+ and recovered with anti-GSTO1-1
(a-G). In d, RyR2 activity increased
with N-ethylmaleimide-treated GSTO1-1 (G-M) and
then declined after adding control-treated GSTO1-1 (G-C).
-CT, control-treated. In contrast to the decrease in
activity of cardiac RyRs exposed to enzymatically active
GSTO1-1, e shows an increase in skeletal RyR1 activity with
GSTO1-1. Continuous lines, zero current; broken
lines, maximum open conductance.
View larger version (38K):
[in a new window]
Fig. 4.
Average effects of cytoplasmic GSTO1-1 on
RyR channels. I't/I'c
is shown, with numbers of bilayers in parentheses (a-e).
a, GSTO1-1 buffer (B) alone did not alter RyR2
activity. a-G, anti-GSTO1-1. b, RyR2 activity
(10 6 M cis
Ca2+) was depressed by GSTO1-1 (G) and enhanced
with anti-GSTO1-1 (channels were re-exposed to 210 µg/ml GSTO1-1
after washout of GSTO1-1 and before exposure to anti-GSTO1-1).
c, GTSO1-1 depressed RyR2 activity with
10
5 M cis
Ca2+. d, RyR2 activity increased with 92 µg/ml
N-ethylmaleimide-treated GSTO1-1-M (G-M) and
then declined with 88 µg/ml control-incubated GSTO1-1-C
(G-C). e, RyR1 activity increased with GSTO1-1.
Asterisks show significant differences between test and
control data. f, average
I't/I'c from four bilayers
exposed sequentially to 2.1, 21, and 210 µg/ml GSTO1-1 plotted
against [GSTO1-1] and fitted with the inhibition equation,
where
I't/I'cmax is
the maximum relative mean current (i.e. 1), EC50
is the concentration of GSTO1-1 required for 50% inhibition, and
B is the fraction of channel activity not inhibited by
GSTO1-1.
(Eq. 1)
Ala mutation that removed enzyme activity
lost its ability to cause depression of RyR2 activity (11 or 110 µg/ml, n = 6). Indeed, RyR2 channel activity was
increased by the mutated GSTO1-1 (n = 4). An increase
in RyR2 activity to levels that were greater than before exposure to
GSTO1-1 after washout of the enzyme was also seen after addition of
antibody and after addition of N-ethylmaleimide-treated enzyme, suggesting that an activating action of GSTO1-1 was unmasked when the inhibitory effect was removed or was not present. Only an
excitatory effect of GSTO1-1 was seen in skeletal muscle RyR1 channels, and this was not reversed within 5 min of removing GSTO1-1 (Figs. 3e and 4e).
DISCUSSION
TOP
ABSTRACT
INTRODUCTION
EXPERIMENTAL PROCEDURES
RESULTS
DISCUSSION
REFERENCES
![]() |
ACKNOWLEDGEMENTS |
---|
We are grateful to Suzi Pace for help with SR vesicle preparation and Marjorie Coggan for help with expression of GSTO.
![]() |
FOOTNOTES |
---|
* The costs of publication of this article were defrayed in part by the payment of page charges. The article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.
To whom correspondence should be addressed. Tel.: 61 2 62494714;
Fax: 61 2 62494712; E-mail: Philip.Board@anu.edu.au.
Published, JBC Papers in Press, October 16, 2000, DOI 10.1074/jbc.M007874200
2 Further details of the alignment are available from Dr. Philip Board at the John Curtin School of Medical Research.
![]() |
ABBREVIATIONS |
---|
The abbreviations used are: GST, glutathione transferase; CLIC, chloride intracellular channel; RyR, ryanodine receptor; SR, sarcoplasmic reticulum; TES, N-tris(hyroxymethyl)methyl-2-aminoethanesulfonic acid.
![]() |
REFERENCES |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
1. | Hayes, J. D., and Pulford, D. J. (1995) Crit. Rev. Biochem. Mol. Biol. 30, 445-600[Abstract] |
2. | Litwack, G., Ketterer, B., and Arias, I. M. (1971) Nature 234, 466-467[Medline] [Order article via Infotrieve] |
3. |
Fernandez-Canon, J. M.,
and Penalva, M. A.
(1998)
J. Biol. Chem.
273,
329-337 |
4. |
Adler, V.,
Yin, Z.,
Fuchs, S. Y.,
Benezra, M.,
Rosario, L.,
Tew, K. D.,
Pincus, M. R.,
Sardana, M.,
Henderson, C. J.,
Wolf, C. R.,
Davis, R. J.,
and Ronai, Z.
(1999)
EMBO J.
18,
1321-1334 |
5. | Board, P. G., Baker, R. T., Chelvanayagam, G., and Jermiin, L. S. (1997) Biochem. J. 328, 929-935[Medline] [Order article via Infotrieve] |
6. | Tong, Z., Board, P. G., and Anders, M. W. (1998) Chem. Res. Toxicol. 11, 1332-1338[CrossRef][Medline] [Order article via Infotrieve] |
7. |
Board, P. G.,
Coggan, M.,
Chelvanayagam, G.,
Easteal, S.,
Jermiin, L. S.,
Schulte, G. K.,
Danley, D. E.,
Hoth, L. R.,
Griffor, M. C.,
Kamath, A. V.,
Rosner, M. H.,
Chrunyk, B. A.,
Perregaux, D. E.,
Gabel, C. A.,
Geoghegan, K. F.,
and Pandit, J.
(2000)
J. Biol. Chem.
275,
24798-24806 |
8. | Wilce, M. C., and Parker, M. W. (1994) Biochim. Biophys. Acta 1205, 1-18[Medline] [Order article via Infotrieve] |
9. |
Kodym, R.,
Calkins, P.,
and Story, M.
(1999)
J. Biol. Chem.
274,
5131-5137 |
10. |
Altschul, S. F.,
Madden, T. L.,
Schaffer, A. A.,
Zhang, J.,
Zhang, Z.,
Miller, W.,
and Lipman, D. J.
(1997)
Nucleic Acids Res.
25,
3389-3402 |
11. |
Valenzuela, S. M.,
Martin, D. K.,
Por, S. B.,
Robbins, J. M.,
Warton, K.,
Bootcov, M. R.,
Schofield, P. R.,
Campbell, T. J.,
and Breit, S. N.
(1997)
J. Biol. Chem.
272,
12575-12582 |
12. |
Qian, Z.,
Okuhara, D.,
Abe, M. K.,
and Rosner, M. R.
(1999)
J. Biol. Chem.
274,
1621-1627 |
13. |
Xu, L.,
Tripathy, A.,
Pasek, D. A.,
and Meissner, G.
(1998)
Ann. N. Y. Acad. Sci.
853,
130-148 |
14. |
Sei, Y.,
Gallagher, K. L.,
and Basile, A. S.
(1999)
J. Biol. Chem.
274,
5995-6002 |
15. | Guse, A. H., da Silva, C. P., Berg, I., Skapenko, A. L., Weber, K., Heyer, P., Hohenegger, M., Ashamu, G. A., Schulze-Koops, H., Potter, B. V., and Mayr, G. W. (1999) Nature 398, 70-73[CrossRef][Medline] [Order article via Infotrieve] |
16. | Mariot, P., Prevarskaya, N., Roudbaraki, M. M., Le Bourhis, X., Van Coppenolle, F., Vanoverberghe, K., and Skryma, R. (2000) Prostate 43, 205-214[CrossRef][Medline] [Order article via Infotrieve] |
17. |
Pan, Z.,
Damron, D.,
Nieminen, A.-L.,
Bhat, M. B.,
and Ma, J.
(2000)
J. Biol. Chem
275,
19978-19984 |
18. | Thompson, J. D., Higgins, D. G., and Gibson, T. J. (1994) Nucleic Acids Res. 22, 4673-4680[Abstract] |
19. | Jones, D. T., Taylor, W. R., and Thornton, J. M. (1992) Nature 358, 86-89[CrossRef][Medline] [Order article via Infotrieve] |
20. | Whittington, A., Vichai, V., Webb, G., Baker, R., Pearson, W., and Board, P. (1999) Biochem. J. 337, 141-151[CrossRef][Medline] [Order article via Infotrieve] |
21. | Laver, D. R., Roden, L. D., Ahern, G. P., Eager, K. R., Junankar, P. R., and Dulhunty, A. F. (1995) J. Membr. Biol. 147, 7-22[Medline] [Order article via Infotrieve] |
22. | Laver, D. R., Owen, V. J., Junankar, P. R., Taske, N. L., Dulhunty, A. F., and Lamb, G. D. (1997) Biophys. J. 73, 1913-1924[Abstract] |
23. | Minium, E. W., King, B. K., and Bear, G. (1993) Statistical Reasoning in Psychology and Education , John Wiley & Sons, Inc., New York |
24. |
Kermode, H.,
Williams, A. J.,
and Sitsapesan, R.
(1998)
Biophys. J.
74,
1296-1304 |
25. |
Dulhunty, A. F.,
Laver, D. R.,
Gallant, E. M.,
Casarotto, M. G.,
Pace, S. M.,
and Curtis, S.
(1999)
Biophys. J.
77,
189-203 |
26. | Dulhunty, A., Haarmann, C., Green, D., and Hart, J. (2000) Antioxidants Redox Signalling 2, 27-34[Medline] [Order article via Infotrieve] |
27. | Villafania, A., Anwar, K., Amar, S., Chie, L., Way, D., Chung, D. L., Adler, V., Ronai, Z., Brandt-Rauf, P. W., Yamaizumii, Z., Kung, H. F., and Pincus, M. R. (2000) Ann. Clin. Lab. Sci. 30, 57-64[Abstract] |
28. | Marx, S. O., Reiken, S., Hisamatsu, Y., Jayaraman, T., Burkhoff, D., Rosemblit, N., and Marks, A. R. (2000) Cell 101, 365-376[Medline] [Order article via Infotrieve] |