ACCELERATED PUBLICATION
Substrate-induced Conformational Changes in the Transmembrane
Segments of Human P-glycoprotein
DIRECT EVIDENCE FOR THE SUBSTRATE-INDUCED FIT MECHANISM FOR DRUG
BINDING*
Tip W.
Loo,
M. Claire
Bartlett, and
David M.
Clarke
From the Canadian Institutes of Health Research Group in Membrane
Biology, Department of Medicine and Department of Biochemistry,
University of Toronto, Toronto, Ontario M5S 1A8, Canada
Received for publication, February 14, 2003, and in revised form, February 26, 2003
 |
ABSTRACT |
The human multidrug resistance P-glycoprotein
(P-gp, ABCB1) is quite promiscuous in that it can transport a broad
range of structurally diverse compounds out of the cell. We
hypothesized that the transmembrane (TM) segments that constitute the
drug-binding site are quite mobile such that drug binding occurs
through a "substrate-induced fit" mechanism. Here, we used
cysteine-scanning mutagenesis and oxidative cross-linking to test for
substrate-induced changes in the TM segments. Pairs of cysteines were
introduced into a Cys-less P-gp and the mutants treated with oxidant
(copper phenanthroline) in the presence or absence of various drug
substrates. We show that cyclosporin A promoted cross-linking between
residues P350C(TM6)/G939C(TM11), while colchicine and demecolcine
promoted cross-linking between residues P350C(TM6)/V991C(TM12).
Progesterone promoted cross-linking between residues
P350C(TM6)/A935C(TM11), P350C(TM6)/G939C(TM11), as well as
between residues P350C(TM6)/V991C(TM12). Other substrates such as
vinblastine, verapamil, cis-(Z)-flupenthixol or
trans-(E)-flupenthixol did not induce
cross-linking at these sites. These results provide direct evidence
that the packing of the TM segments in the drug-binding site is changed
when P-gp binds to a particular substrate. The induced-fit mechanism
explains how P-gp can accommodate a broad range of compounds.
 |
INTRODUCTION |
P-glycoprotein (P-gp)1
is an ATP-dependent drug pump that is capable of
transporting structurally diverse compounds out of the cell (recently
reviewed in Refs. 1 and 2). P-gp is a member of the
ATP-Binding Cassette (ABC) family
of transporters. The 1280 amino acids of P-gp are arranged as two
repeating units of 610 amino acids that are joined by a linker region
of about 60 amino acids (3). Each repeat has six transmembrane (TM) segments and a hydrophilic domain containing an ATP-binding site (4-6). P-gp functions as a monomer (7), but the two halves of the
molecule do not have to be covalently linked for function (8, 9).
Studies on deletion mutants have shown that the TM domains alone are
sufficient to mediate drug binding (9). Drug binding requires the
contribution of the six NH2-terminal TM segments and the
six COOH-terminal TM segments (10).
An important goal in understanding the mechanism of P-gp is to
determine how P-gp recognizes so many structurally diverse compounds.
Studies on cysteine mutants and their reactivity with different
thiol-reactive substrate analogs indicate that the drug-binding site is
lined with residues from multiple TM segments (11-16). Therefore, it
has been proposed that P-gp binds these substrates through an
induced-fit mechanism where the size and shape of the substrate changes
packing of the TM segments.
In this study we used cysteine-scanning mutagenesis and oxidative
cross-linking to test for rearrangement of the TM segments in the
presence of different substrates.
 |
MATERIALS AND METHODS |
Construction of Mutants--
A histidine-tagged Cys-less P-gp
cDNA was constructed (4, 17). Pairs of cysteines were then
introduced into the Cys-less P-gp (18). The presence of the histidine
tag facilitated purification of the mutant P-gps by nickel-chelate
chromatography (19). A cDNA coding for the TM domains only (TMD1+2,
residues 1-379 + residues 681-1025) and tagged with the epitope for
monoclonal antibody A52 was constructed as described previously (9,
20).
Expression and Disulfide Cross-linking Analysis--
HEK 293 cells were transfected with the mutant cDNAs. After 24 h, the
medium was replaced with fresh media and the cells grown for
72 h at 27 °C. Membranes were prepared as described previously (17, 19) and used for disulfide cross-linking analysis (21) by
incubation with Cu2+(phenanthroline)3 (0.2 mM final concentration) under the conditions described in
the figure legends.
Expression, Purification, and Measurement of Drug-stimulated
ATPase Activity of P-gp Mutants--
The histidine-tagged P-gp mutants
were expressed in HEK 293 cells, purified, and mixed with lipid as
described previously (19). An aliquot of the P-gp:lipid mixture was
assayed for drug-stimulated ATPase activity by addition of an equal
volume of buffer containing 100 mM Tris-HCl, pH 7.5, 100 mM NaCl, 20 mM MgCl2, 10 mM ATP and 10 mM colchicine, 2 mM
demecolcine, 0.4 mM progesterone, 0.2 mM cis-(Z)-flupenthixol, 1 mm verapamil, or 20 µM vinblastine. These concentrations gave maximal
stimulation of the mutant P-gp mutant P350C. The samples were incubated
for 30 min at 37 °C, and the amount of inorganic phosphate liberated
was determined (22).
 |
RESULTS |
We had proposed that substrate binding occurred through a
"substrate-induced fit" mechanism (14). The TM segments
contributing residues to the drug-binding site are predicted to be
quite mobile so that drug binding causes rearrangement of the TM
segments. Cysteine-scanning mutagenesis and oxidative cross-linking
studies are ideal techniques for detecting altered packing of the TM
segments in the presence of drug substrates. The TM segments in the
resting state are arranged in a "funnel shape" in which the
cytoplasmic ends are close together (15, 21). The cytoplasmic side of TM6 and TM12 is relatively close together, because mutant
P350C(TM6)/S993C(TM12) could be cross-linked. We showed that
cross-linking of this mutant inhibited drug-stimulated ATPase activity,
but activity was restored after addition of a thiol reducing agent
(23). Therefore, it may be possible to detect changes in the
cross-linking pattern if P350C(TM6) or S993C(TM12) is paired with
another cysteine at the cytoplasmic end of other TM segments.
Accordingly, 143 double cysteine mutants that were
constructed previously were used in this study (21). These mutants
contained P350C(TM6) and another cysteine on the cytoplasmic half of
TMs 7-12 (i.e. P350C + another cysteine at positions:
711-723 (TM7), 770-783 (TM8), 828-840 (TM9), 867-879 (TM10),
935-947 (TM11), and 986-994 (TM12)) or S993C(TM12) with another
cysteine in the cytoplasmic half of TMs 1-6 (i.e. S993C + another cysteine at positions: 51-61 (TM1), 130-141 (TM2), 185-196
(TM3), 226-237 (TM4), 293-304 (TM5), and 343-351 (TM6)). We had
shown that most of the double cysteine mutants retained at least 70%
of the verapamil-stimulated ATPase activity of the Cys-less P-gp (21).
We then tested compounds of different shapes and sizes (colchicine,
demecolcine, verapamil, vinblastine, cyclosporin A, progesterone,
cis-(Z)-flupenthixol and
trans-(E)-flupenthixol) for their ability to
promote cross-linking. Vinblastine and colchicine are classic transport
substrates of P-gp (24, 25). Demecolcine and verapamil induce large
increases in ATPase activity of P-gp (26). Cyclosporin A is a large
cyclic peptide substrate (27), while the flupenthixol isomers have opposite effects on the ATPase activity of P-gp (28). Progesterone is a
steroid substrate of P-gp (29, 30). The mutants were expressed
transiently in HEK 293 cells. Membranes were prepared and preincubated
at 21 °C for 10 min with or without saturating levels of colchicine
(5 mM), demecolcine (1 mM), verapamil (1 mM), vinblastine (0.1 mM), cyclosporin A (0.1 mM), cis-(Z)-flupenthixol (2 mM), trans-(E)-flupenthixol (2 mM), or progesterone (2 mM). The membranes were
treated with 0.2 mM copper phenanthroline (oxidant) for 10 min at 21 °C. The reactions were stopped by addition of EDTA and the
samples subjected to SDS-PAGE and immunoblot analysis. Fig.
1 shows that in the absence of oxidant,
mature P-gp migrates with an apparent mass of 170 kDa. Cross-linked
P-gp, however, migrates with a slower mobility in SDS-PAGE (18, 31,
32). In most mutants, cross-linked product was not detected in the presence of drug substrates. An example is mutant
P350C(TM6)/A869C(TM10) (Fig. 1). Three mutants (P350C(TM6)/A935C(TM11),
P350C(TM6)/G939C(TM11) and P350C(TM6)/V991C(TM12)), however, showed
drug-induced cross-linking (Fig. 1). Mutant P350C(TM6)/A935C(TM11) was
cross-linked only in the presence of progesterone. Cross-linking of
mutant P350C(TM6)/G939C(TM11) was enhanced in the presence of
cyclosporin A and progesterone, while mutant P350C(TM6)/V991C(TM12)
showed enhanced cross-linking with colchicine, demecolcine, and
progesterone (Fig. 1).

View larger version (59K):
[in this window]
[in a new window]
|
Fig. 1.
Effect of drugs on disulfide cross-linking of
P-gp mutants. Membranes were prepared from HEK 293 cells
expressing mutants P350C(TM6)/A935C(TM11),
P350C(TM6)/G939C(TM11), P350C(TM6)/V991C(TM12), and
P350C(TM6)/A869C(TM10). The membranes were incubated at 21 °C for 10 min with 5 mM colchicine (Colch), 1 mM demecolcine (Dem), 1 mM verapamil
(Ver), 0.1 mM vinblastine (Vin), 0.1 mM cyclosporin A (Cyclo), 2 mM
cis-(Z)-flupenthixol (Cis-Flu), 2 mM trans-(E)-flupenthixol
(Trans-Flu), 2 mM progesterone
(Prog), or no drug substrate. The reaction mixtures were
then treated with oxidant for 10 min at 21 °C with 0.2 mM copper phenanthroline (oxidant). The
reactions were stopped by addition of SDS sample buffer containing 50 mM EDTA and no reducing agent. The mixtures were subjected
to immunoblot analysis. The positions of the cross-linked
(X-link) product and mature (170 kDa) P-gp are
indicated.
|
|
It was important to express the mutant P-gps in the absence of drug
substrate or to thoroughly wash the membranes several times with
tris-buffered saline if they were prepared from cells that had been
exposed to a drug substrate. This is because residual substrate in the
membrane could potentially influence the cross-linking pattern. An
example was mutant P350C(TM6)/G939C(TM11). When this mutant was
expressed in the absence of cyclosporin A, it showed no cross-linking
at 21 °C (Fig. 1), but some cross-linking was detected when
membranes prepared from cyclosporin A-treated cells were treated with
oxidant (21). Therefore, in this study, membranes were prepared from
transfected cells that were not treated with any substrate.
To compare the effectiveness of the drug substrates in
promoting cross-linking, time-dependent cross-linking
studies were done on P350C(TM6)/A935C(TM11), P350C(TM6)/G939C(TM11),
and P350C(TM6)/V991C(TM12). Fig. 2 shows
that in the absence of drug substrate, no cross-linked product was
detected in any of the mutants. Progesterone, however, promoted
cross-linking in all three mutants and was the only drug substrate that
promoted cross-linking of mutant P350C(TM6)/A935C(TM11). In mutant
P350C(TM6)/G939C(TM11), progesterone was more efficient in
promoting cross-linking than cyclosporin A. In mutant
P350C(TM6)/V991C(TM12), the drug substrates colchicine,
demecolcine, and progesterone were equally effective in promoting
cross-linking.

View larger version (63K):
[in this window]
[in a new window]
|
Fig. 2.
Time-dependent cross-linking of
P-gp mutants. Membranes prepared from HEK 293 cells expressing
mutants P350C(TM6)/A935C(TM11), P350C(TM6)/G939C(TM11), or
P350C(TM6)/V991C(TM12) were preincubated for 10 min at 21 °C
with no drug, 1 mM progesterone, 0.1 mM
cyclosporin A, 5 mM colchicine, or 1 mM
demecolcine. The reaction mixtures were then treated with 0.2 mM copper phenanthroline at 21 °C for the indicated
times. The reaction was stopped by addition of SDS sample buffer
containing 50 mM EDTA and no reducing agent. The mixtures
were subjected to immunoblot analysis. The positions of the
cross-linked (X-link) product and mature (170 kDa) P-gp are
indicated.
|
|
Cross-linking promoted by the presence of drug substrates cyclosporin
A, colchicine, demecolcine, or progesterone could be explained on the
basis that drug binding to the TM domain alters TM segment packing. It
has been reported, however, that some drug substrates such as
progesterone bind to mouse P-gp in a region of the nucleotide-binding
domain that is in close proximity to the ATP site (33). To test whether
cyclosporin A, colchicine, demecolcine, or progesterone can bind to the
TM domains, we used a "drug rescue" assay involving a P-gp mutant
that lacked the NBDs (TMD1+2, residues 1-379 + residues 681-1025).
The rationale for the drug rescue assay is that the TMD1+2 mutant is
misfolded when transiently expressed in the absence of drug substrate
and is retained within the cell as a 80-kDa core-glycosylated protein (34). Expression of the mutant TMD1+2 in the presence of drug substrate, however, induces the mutant protein to fold properly into a
100-kDa protein endoglycosidase H-resistant form that is transported to
the cell surface (9). It appears that the drug substrate diffuses into
the cell and acts as a specific chemical chaperone to bind and
stabilize the newly synthesized misfolded P-gp that is present
transiently and thereby induce proper folding and trafficking of the
protein (10, 35).
The TMD1+2 deletion mutant was expressed in HEK 293 cells with or
without demecolcine, colchicine, or progesterone. The cells were
solubilized with SDS buffer and subjected to immunoblot analysis. Fig.
3 shows that mutant TMD1+2 is expressed
as an 80-kDa protein in the absence of substrate. In the presence of
demecolcine, colchicine, or progesterone, however, the presence of a
100-kDa protein is detected. Cyclosporin A also induced proper folding
of mutant TMD1+2 (9). Therefore, these drug substrates can interact
with only the TM domains.

View larger version (46K):
[in this window]
[in a new window]
|
Fig. 3.
Effect of drug substrates on expression of
P-gp mutant TMD1+2. HEK 293 cells were transfected with the
A52-tagged P-gp mutant TMD1+2 (residues 1-379 + residues 681-1025)
cDNA. After 24 h, the cells were incubated for another 24 h with fresh medium containing no drug substrate (No
drug) or with 1 mM colchicine (Colch), 0.5 mM demecolcine (Dem), or 0.1 mM
progesterone (Prog). The cells were then solubilized with
SDS sample buffer and samples subjected to immunoblot analysis. The
positions of the mature (100 kDa) and immature (80 kDa) proteins are
indicated.
|
|
We then tested whether the mutants could interact with colchicine,
demecolcine, progesterone, cis-(Z)-flupenthixol,
verapamil, and vinblastine, because only some of the drugs affected the
cross-linking pattern of the mutants. It was important to determine
that the drugs that had no effect on cross-linking could still bind to the mutant P-gps. There is good correlation between drug-stimulated ATPase activity and drug transport (36) (37). Accordingly, we tested
whether the drug substrates colchicine, demecolcine, progesterone,
cis-(Z)-flupenthixol, verapamil, or vinblastine stimulated the ATPase activities of histidine-tagged mutants
P350C(TM6)/A935C(TM11), P350C(TM6)/G939C(TM11), and
P350C(TM6)/V991C(TM12). Their activities were compared with that
of mutant P350C (parent). We previously showed that mutant P350C
exhibited high levels of drug-stimulated ATPase activity (11). Fig.
4 shows that the ATPase activities of
mutants P350C(TM6)/A935C(TM11), P350C(TM6)/G939C(TM11), and P350C(TM6)/V991C(TM12) were stimulated by colchicine, demecolcine, progesterone, cis-(Z)-flupenthixol, verapamil, or
vinblastine. The ATPase activities of the mutants were inhibited by
cyclosporin A and trans-(Z)-flupenthixol (data
not shown). This is consistent with previous observations that
saturating concentrations of cyclosporin A inhibits P-gp ATPase
activity, while the cis-(Z)- and
trans-(E)-isomers of flupenthixol stimulate and
inhibit, respectively, the ATPase activity of P-gp (26, 38, 39). These
results indicate that the mutant P-gps can still bind drug
substrates.

View larger version (19K):
[in this window]
[in a new window]
|
Fig. 4.
Relative drug-stimulated ATPase activities of
P-gp mutants. Histidine-tagged P-gp mutants
P350C(TM6)/A935C(TM11), P350C(TM6)/G939C(TM11), or
P350C(TM6)/V991C(TM12) were expressed in HEK 293 cells and isolated by
nickel-chelate chromatography. The isolated proteins were mixed with
lipid, sonicated, and drug-stimulated ATPase activity measured in the
presence of 5 mM colchicine (Colch), 1 mM demecolcine (Dem), 0.2 mM
progesterone (Prog), 0.1 mM
cis-(Z)-flupenthixol (Cis-Flu), 0.5 mM verapamil (Ver), or 10 µM
vinblastine (Vin). The activities of the mutant P-gps are
expressed relative to mutant P350C.
|
|
 |
DISCUSSION |
Cross-linking of mutant P350C(TM6)/S993(TM12) is inhibited by drug
substrates (23). In the present study, we show that substrates such as
progesterone are effective in promoting new cross-links with P350C
(Fig. 2). In the absence of drug substrate, residue P350C in TM6 can be
cross-linked to S993C in TM12. The presence of progesterone, however,
promoted cross-linking of residue P350C(TM6) with two residues in TM 11 (A935C and G939C) and to residue V991C in TM12. One way to explain
these results is through a model (Fig. 5). Based on the "funnel-shape" model
of the TM segments (15), the cytoplasmic ends of TMs 6, 11, and 12 are
placed closer together than the extracellular ends (Fig.
5A). In the absence of substrate, it is possible to
cross-link P350C(TM6)/S993(TM12). Therefore, the presence of drug
substrate (progesterone) likely causes a slight rotation/rearrangement
of TM12 relative to TM6 such that residue V991C comes closer to P350C.
Colchicine and demecolcine likely induced similar conformational
changes, since both drug substrates also promoted cross-linking of
mutant P350C(TM6)/V991C(TM12). Since TM12 is directly connected to
TM11, any (rotational or lateral) movement in TM12 would likely involve
similar movement in TM11. When the residues in TM11 are modeled as an
-helical wheel, residues A935 and G939C are found on the same face
of the TM segment (Fig. 5B). In the presence of progesterone
both residues must come close to P350C(TM6) to be cross-linked (Fig.
1).

View larger version (41K):
[in this window]
[in a new window]
|
Fig. 5.
Model of progesterone-induced conformational
changes in the TM segments. A, cylinders
represent TM segments 6, 11, and 12. In the absence of drug substrate,
disulfide cross-linking occurs between Pro350
(yellow ball) and S993C (purple ball). In the
presence of drug substrate (progesterone), TM segments 11 and 12 undergo rotational and/or lateral movements so that cross-linking can
occur between P350C and V991C (black ball) in TM12 and with
A935C (red ball) and G939C (turquoise ball) in
TM11. B, residues in TM11 are modeled in an -helical
wheel. Residues Ala935 and Gly939 appear on the
same face of the helix.
|
|
Cyclosporin A also promoted cross-linking between TM6 and TM11, but
only between P350C and G939C. It is possible that cyclosporin A altered
the tilt or distance between TM6 and TM11 such that only A939C but not
A935C were close enough to P350C to be cross-linked.
The ability of a substrate to change the cross-linking pattern suggests
that the TM segments can change their shape to accommodate structurally
different compounds. Slight rotational and/or lateral movement in any
TM segment could result in numerous permutations of residues
contributing to the drug-binding site. A substrate with one structure
would cause specific shifts in the different TM segments responsible
for its binding (induced-fit). Therefore, it follows that common
residues could be involved in the binding of different substrates. This
would account for the ability of P-gp to bind structurally diverse compounds.
 |
FOOTNOTES |
*
This work was supported in part by United States National
Institutes of Health Grant CA80900 and by grants from the Canadian Institutes for Health Research and the Canadian Cystic Fibrosis Foundation.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.
Recipient of the Canada Research Chair in Membrane Biology. To
whom correspondence should be addressed: Dept. of Medicine, University
of Toronto, Rm. 7342, Medical Sciences Bldg., 1 King's College Circle,
Toronto, Ontario M5S 1A8, Canada. Tel.:/Fax: 416-978-1105; E-mail:
david.clarke@utoronto.ca.
Published, JBC Papers in Press, February 27, 2003, DOI 10.1074/jbc.C300073200
 |
ABBREVIATIONS |
The abbreviations used are:
P-gp, P-glycoprotein;
TM, transmembrane;
HEK, human embryonic kidney;
TMD1+2, residues 1-379 + residues 681-1025 of P-gp.
 |
REFERENCES |
1.
|
Ambudkar, S. V.,
Dey, S.,
Hrycyna, C. A.,
Ramachandra, M.,
Pastan, I.,
and Gottesman, M. M.
(1999)
Annu. Rev. Pharmacol. Toxicol.
39,
361-398[CrossRef][Medline]
[Order article via Infotrieve]
|
2.
|
Borst, P.,
and Elferink, R. O.
(2002)
Annu. Rev. Biochem.
71,
537-592[CrossRef][Medline]
[Order article via Infotrieve]
|
3.
|
Chen, C. J.,
Chin, J. E.,
Ueda, K.,
Clark, D. P.,
Pastan, I.,
Gottesman, M. M.,
and Roninson, I. B.
(1986)
Cell
47,
381-389[Medline]
[Order article via Infotrieve]
|
4.
|
Loo, T. W.,
and Clarke, D. M.
(1995)
J. Biol. Chem.
270,
843-848[Abstract/Free Full Text]
|
5.
|
Loo, T. W.,
and Clarke, D. M.
(1995)
J. Biol. Chem.
270,
22957-22961[Abstract/Free Full Text]
|
6.
|
Kast, C.,
Canfield, V.,
Levenson, R.,
and Gros, P.
(1995)
Biochemistry
34,
4402-4411[Medline]
[Order article via Infotrieve]
|
7.
|
Loo, T. W.,
and Clarke, D. M.
(1996)
J. Biol. Chem.
271,
27488-27492[Abstract/Free Full Text]
|
8.
|
Loo, T. W.,
and Clarke, D. M.
(1994)
J. Biol. Chem.
269,
7750-7755[Abstract/Free Full Text]
|
9.
|
Loo, T. W.,
and Clarke, D. M.
(1999)
J. Biol. Chem.
274,
24759-24765[Abstract/Free Full Text]
|
10.
|
Loo, T. W.,
and Clarke, D. M.
(1998)
J. Biol. Chem.
273,
14671-14674[Abstract/Free Full Text]
|
11.
|
Loo, T. W.,
and Clarke, D. M.
(1997)
J. Biol. Chem.
272,
31945-31948[Abstract/Free Full Text]
|
12.
|
Loo, T. W.,
and Clarke, D. M.
(1999)
J. Biol. Chem.
274,
35388-35392[Abstract/Free Full Text]
|
13.
|
Loo, T. W.,
and Clarke, D. M.
(2000)
J. Biol. Chem.
275,
39272-39278[Abstract/Free Full Text]
|
14.
|
Loo, T. W.,
and Clarke, D. M.
(2001)
J. Biol. Chem.
276,
14972-14979[Abstract/Free Full Text]
|
15.
|
Loo, T. W.,
and Clarke, D. M.
(2001)
J. Biol. Chem.
276,
36877-36880[Abstract/Free Full Text]
|
16.
|
Loo, T. W.,
and Clarke, D. M.
(2002)
J. Biol. Chem.
277,
44332-44338[Abstract/Free Full Text]
|
17.
|
Loo, T. W.,
and Clarke, D. M.
(1993)
J. Biol. Chem.
268,
19965-19972[Abstract/Free Full Text]
|
18.
|
Loo, T. W.,
and Clarke, D. M.
(1996)
J. Biol. Chem.
271,
27482-27487[Abstract/Free Full Text]
|
19.
|
Loo, T. W.,
and Clarke, D. M.
(1995)
J. Biol. Chem.
270,
21449-21452[Abstract/Free Full Text]
|
20.
|
Loo, T. W.,
and Clarke, D. M.
(1993)
J. Biol. Chem.
268,
3143-3149[Abstract/Free Full Text]
|
21.
|
Loo, T. W.,
and Clarke, D. M.
(2000)
J. Biol. Chem.
275,
5253-5256[Abstract/Free Full Text]
|
22.
|
Chifflet, S.,
Torriglia, A.,
Chiesa, R.,
and Tolosa, S.
(1988)
Anal. Biochem.
168,
1-4[Medline]
[Order article via Infotrieve]
|
23.
|
Loo, T. W.,
and Clarke, D. M.
(1997)
J. Biol. Chem.
272,
20986-20989[Abstract/Free Full Text]
|
24.
|
Horio, M.,
Chin, K. V.,
Currier, S. J.,
Goldenberg, S.,
Williams, C.,
Pastan, I.,
Gottesman, M. M.,
and Handler, J.
(1989)
J. Biol. Chem.
264,
14880-14884[Abstract/Free Full Text]
|
25.
|
Choi, K.,
Frommel, T. O.,
Stern, R. K.,
Perez, C. F.,
Kriegler, M.,
Tsuruo, T.,
and Roninson, I. B.
(1991)
Proc. Natl. Acad. Sci. U. S. A.
88,
7386-7390[Abstract]
|
26.
|
Loo, T. W.,
Bartlett, M. C.,
and Clarke, D. M.
(2003)
J. Biol. Chem.
278,
1575-1578[Abstract/Free Full Text]
|
27.
|
Tamai, I.,
and Safa, A. R.
(1991)
J. Biol. Chem.
266,
16796-16800[Abstract/Free Full Text]
|
28.
|
Dey, S.,
Hafkemeyer, P.,
Pastan, I.,
and Gottesman, M. M.
(1999)
Biochemistry
38,
6630-6693[CrossRef][Medline]
[Order article via Infotrieve]
|
29.
|
Yang, C. P.,
Cohen, D.,
Greenberger, L. M.,
Hsu, S. I.,
and Horwitz, S. B.
(1990)
J. Biol. Chem.
265,
10282-10288[Abstract/Free Full Text]
|
30.
|
Vo, Q. D.,
and Gruol, D. J.
(1999)
J. Biol. Chem.
274,
20318-20327[Abstract/Free Full Text]
|
31.
|
Loo, T. W.,
and Clarke, D. M.
(2000)
J. Biol. Chem.
275,
19435-19438[Abstract/Free Full Text]
|
32.
|
Urbatsch, I. L.,
Gimi, K.,
Wilke-Mounts, S.,
Lerner-Marmarosh, N.,
Rousseau, M. E.,
Gros, P.,
and Senior, A. E.
(2001)
J. Biol. Chem.
276,
26980-26987[Abstract/Free Full Text]
|
33.
|
Dayan, G.,
Baubichon-Cortay, H.,
Jault, J. M.,
Cortay, J. C.,
Deleage, G.,
and Di Pietro, A.
(1996)
J. Biol. Chem.
271,
11652-11658[Abstract/Free Full Text]
|
34.
|
Loo, T. W.,
and Clarke, D. M.
(1994)
J. Biol. Chem.
269,
28683-28689[Abstract/Free Full Text]
|
35.
|
Loo, T. W.,
and Clarke, D. M.
(1997)
J. Biol. Chem.
272,
709-712[Abstract/Free Full Text]
|
36.
|
Ambudkar, S. V.,
Cardarelli, C. O.,
Pashinsky, I.,
and Stein, W. D.
(1997)
J. Biol. Chem.
272,
21160-21166[Abstract/Free Full Text]
|
37.
|
Doige, C. A., Yu, X.,
and Sharom, F. J.
(1992)
Biochim. Biophys. Acta
1109,
149-160[Medline]
[Order article via Infotrieve]
|
38.
|
Sarkadi, B.,
and Muller, M.
(1997)
Semin. Cancer Biol.
8,
171-182[CrossRef][Medline]
[Order article via Infotrieve]
|
39.
|
Dey, S.,
Ramachandra, M.,
Pastan, I.,
Gottesman, M. M.,
and Ambudkar, S. V.
(1997)
Proc. Natl. Acad. Sci. U. S. A.
94,
10594-10599[Abstract/Free Full Text]
|
Copyright © 2003 by The American Society for Biochemistry and Molecular Biology, Inc.