From the Department of Molecular Pharmacology and
Toxicology, School of Pharmacy and the § Department of Cell
and Neurobiology, School of Medicine, University of Southern
California, Los Angeles, California 90089
Received for publication, August 2, 2000, and in revised form, December 26, 2000
![]() |
ABSTRACT |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
Monoamine oxidase (MAO) is responsible for the
oxidation of biogenic and dietary amines. It exists as two isoforms, A
and B, which have a 70% amino acid identity and different substrate and inhibitor specificities. This study reports the identification of
residues responsible for conferring this specificity in human MAO A and
B. Using site-directed mutagenesis we reciprocally interchanged three pairs of corresponding nonconserved amino acids within the central portion of human MAO. Mutant MAO A-I335Y became like MAO B,
which exhibits a higher preference for Monoamine oxidase (MAO1;
amine:oxygen oxidoreductase (deaminating) (flavin-containing), EC
1.4.3.4) catalyzes the oxidative deamination of biogenic and xenobiotic
amines and plays an important role in regulating their levels. MAO is a
flavin-adenine dinucleotide-containing enzyme located on the
mitochondrial outer membrane (1-3). It exists in two forms, A and B. MAO A preferentially oxidizes serotonin (5-hydroxytryptamine, 5-HT) and
is inhibited by low concentrations of clorgyline (4) and Ro 41-1049
(5), whereas MAO B preferentially oxidizes To determine the region(s) responsible for the substrate and inhibitor
specificities of the two isoenzymes, we and other groups have made
point mutations and chimeric MAO A/B enzymes (15-21). It has been
shown that reciprocally interchanging amino acids Phe-208 in MAO A and
its corresponding residue in MAO B, Ile-199, was sufficient to
partially reverse the substrate and inhibitor specificities of rat MAOs
(22) but not human MAOs (21). This indicated that different amino acid
residues may determine specificity in human and rat MAOs. We also found
that the residues that may be important for specificity in human MAOs
are within a 161-amino acid segment (amino acids 215-375 in MAO A and
206-266 in MAO B) (21). This segment contains 32 amino acids that are
nonconserved between human MAO A and B, of which 15 are conserved among
the different species of MAO A or B. Trout MAO shares a 70 and 71% amino acid identity with MAO A and B, respectively. However because the
substrate and inhibition profile of trout MAO is much closer to that of
MAO A than to that of MAO B (23), classifying it as such allows us to
reduce the number of amino acids potentially responsible for
specificity from 15 to 5 (Fig. 1). Of
these five, two were mutated as part of an earlier experiment to the
corresponding residue in the other MAO, expressed in
Saccharomyces cerevisiae cells, and did not exhibit
any kinetic differences from the wild-type enzyme.2 This indicated that
they do not play an important role in specificity. For the other three
amino acids we have made six reciprocal mutants. We found that when two
corresponding amino acids, Ile-335 and Tyr-326 in human MAO A and B,
respectively, were reciprocally interchanged, the substrate and
inhibitor specificities were also switched. This result suggests that
Ile-335 and Tyr-326 in human MAO A and B, respectively, play a key role
in conferring substrate and inhibitor specificities in human
MAOs.
Materials--
The QuikChangeTM site-directed mutagenesis kit
was purchased from Stratagene (La Jolla, CA). The oligonucleotides were
from Operon Technologies Inc. (Alameda, CA). 5-HT, PEA, clorgyline, and
( Construction of the Baculovirus Transfer Vector--
Full-length
human MAO A and B cDNAs were subcloned into the EcoRI
site of pVL1392 transfer vector and named pVL1392-hMAOA and
pVL1392-hMAOB, respectively. The clone was verified for proper insertion by restriction digest analysis. Recombinant MAO A and B-encoding virus was produced by homologous recombination and cotransfection of Sf21 insect cells with the transfer vector and the linearized baculovirus DNA via the calcium phosphate method (24).
Recombinant baculovirus was isolated by plaque purification and
amplified by infection of insect cells with a multiplicity of infection
of 1. After four passes a viral stock with a titer of 108
plaque-forming units/ml was produced.
Site-directed Mutagenesis of Human MAO A and
B--
Site-directed mutagenesis on pVL1392-hMAOA and pVL1392-hMAOB
were performed using the QuikChange kit and following the protocol of
the kit. The following complementary 5'-primers were used: A-T245I, GAACCATCCTGTCATTCACGTTGACC,
GGTCAACGTGAATGACAGGATGGTTC; A-D328G,
GCATGATCATTGAAGGTGAAGATGCTCC,
GGAGCATCTTCACCTTCAATGATCATGC; A-I335Y,
GCTCCAATTTCATACACCTTGGATGAC,
GTCATCCAAGGTGTATGAAATTGGAGC; B-I236T,
GGAGAGGCCTGTGACCTACATTGACC,
GGTCAATGTAGGTCACAGGCCTCTCC; B-G319D,
CCATGATTATTGATGATGAAGAAGCTCC, GGAGCTTCTTCATCATCAATAATCATGG; B-Y326I,
CCAGTTGCCATCACGTTGGATG, CATCCAACGTGATGGCAACTGG; A-I335F, GCTCCAATTTCATTCACCTTGGATGAC,
GTCATCCAAGGTGAATGAAATTGGAGC; B-Y326V, CCAGTTGCCGTCACGTTGGATG, CATCCAACGTGACGGCAACTGG.
The mutagenic bases are underlined. The introduction of the
mutations was confirmed by dideoxy sequencing analysis of the
region of interest.
Expression of the Mutant Clones--
The wild-type and mutant
MAOs were overexpressed in adherent cultures of Sf21 insect
cells (24). 150-mm cell culture dishes were seeded with 20 × 106 Sf21 cells, and recombinant virus was added at a
multiplicity of infection of 2. The cells were incubated at 27 °C
for 72-80 h and harvested by centrifugation for 10 min at 5000 × g. Cell pellets were homogenized in 5 ml of 20 mM sodium phosphate buffer, pH 7.4 containing 0.5 mM EDTA and 0.5 mM phenylmethylsulfonyl fluoride using the polytron homogenizer for 15 s at setting 5. The
homogenates were divided into 0.5-ml aliquots, frozen by dipping into
liquid nitrogen, and stored at Determination of Enzyme Concentration--
The homogenates were
separated by SDS-polyacrylamide gel electrophoresis (25) and stained by
Coomassie Brilliant Blue R-250. A dense band appeared at the expected
molecular mass of MAO (~60 kDa) and represented >70% of the
total protein. The identity of the band was further confirmed by
Western analysis using rabbit anti-MAO A and anti-MAO B antibodies
coupled to hydrogen peroxidase. Signals were developed using a
peroxidase-conjugated goat IgG as secondary antibody followed by
exposure to 3,3'-diaminobenzidine and H2O2. A
highly purified bovine serum albumin standard was electrophoresed in
parallel with the homogenates at nine concentration points ranging from
0.1 to 8 µM. The densities of the MAOs and the bovine
serum albumin bands were quantitated using the EagleSight software, and
the MAO concentrations were calculated according to the linear bovine
serum albumin standard.
Determination of the Kinetic Constants--
The kinetic
constants for the oxidation of 5-HT and PEA and the inhibition by
clorgyline, deprenyl, Ro 41-1040, and Ro 16-6491 were determined by
the radiochemical method as previously described (26) using
O2-saturated 50 mM sodium phosphate buffer. For
the Km determination [14C]5-HT and
[14C]PEA concentrations ranged from 0.1 to 5 times the
Km values that were determined via an Eadie-Hofstee
plot (v versus v/[S]). The
kcat values were calculated from the
Vmax values obtained by fitting the [S]
versus activity curve to the Michaelis-Menten equation and
the calculated concentration of the enzyme from the quantitation assay.
The IC50 values for the irreversible inhibitors clorgyline
and deprenyl were determined by preincubating the inhibitor with the
homogenate for 30 min at 37 °C and assaying for the remaining activity as described (26). The reversible inhibitors were incubated with PEA without the 30-min preincubation. Inhibitor concentrations ranged from 10 We reciprocally interchanged amino acids Thr-245, Asp-328, and
Ile-335 individually in human MAO A with their corresponding amino
acids in human MAO B to produce the mutants A-T245I, A-D328G, and
A-I335Y. Their equivalent mutants in MAO B (B-I236T, B-G319D, and
B-Y326I) were also made.
Wild-type MAO A and B, as well as the six mutants, were successfully
overexpressed in insect cells using recombinant baculovirus. The
turnover number (kcat) and the affinity
(Km) toward the MAO A-specific substrate 5-HT and
the MAO B-specific substrate PEA were determined. We used the
specificity constant,
kcat/Km, to depict the
specificity of an enzyme toward 5-HT or PEA.
As shown in Table I, MAO A wild type had
a 6-fold higher kcat for 5-HT than for PEA but a
similar Km for both substrates, resulting in a
kcat/Km value for 5-HT that
is about seven times that of PEA (Fig.
2). MAO B wild type, on the other hand, had a 19-fold higher kcat for PEA than for 5-HT
and a much lower Km for PEA, resulting in a
kcat/Km for PEA that is about
40,000 times that for 5-HT. Similarly, MAO A and B had a higher
kcat/Km for 5-HT and PEA
respectively. These results are consistent with literature findings
that classify 5-HT as MAO A-specific and PEA as MAO B-specific (16, 17, 26).
-phenylethylamine than for the MAO A preferred substrate serotonin (5-hydroxytryptamine), and
became more sensitive to deprenyl (MAO B-specific inhibitor) than to
clorgyline (MAO A-specific inhibitor). The reciprocal mutant MAO
B-Y326I exhibited an increased preference for 5-hydroxytryptamine, a
decreased preference for
-phenylethylamine, and, similar to MAO A,
was more sensitive to clorgyline than to deprenyl. These mutants also
showed a distinct shift in sensitivity for the MAO A- and B-selective
inhibitors Ro 41-1049 and Ro 16-6491. Mutant pair MAO A-T245I and MAO
B-I236T and mutant pair MAO A-D328G and MAO B-G319D reduced catalytic
activity but did not alter specificity. Our results indicate that
Ile-335 in MAO A and Tyr-326 in MAO B play a critical role in
determining substrate and inhibitor specificities in human MAO A and B.
INTRODUCTION
TOP
ABSTRACT
INTRODUCTION
EXPERIMENTAL PROCEDURES
RESULTS
DISCUSSION
REFERENCES
-phenylethylamine (PEA)
and is inhibited by low concentrations of (
)-deprenyl (6) and Ro
16-6491 (7). Dopamine is a common substrate (8). MAO A and B are
composed of 527 and 520 amino acids, respectively, and have a 70%
amino acid identity (9). They are closely linked on the X-chromosome
(10) and have an identical intron-exon organization, indicating that
they are derived from a common ancestral gene (11). Higher 5-HT and
norepinephrine levels and a phenotype characterized by increased
aggressive behavior is observed when the MAO A gene is deficient in
humans (12) and in mice (13). Disruption of the MAO B gene in mice
results in increased PEA but not 5-HT, norepinephrine, or
dopmanie and confers a resistance to the Parkinsonism-inducing toxin
1-methyl-4phenyl-1,2,3,6-tetrahydropyridine (14). Because MAO is
an integral membrane protein, it is difficult to crystallize, and its
three-dimensional structure has not been reported.
View larger version (59K):
[in a new window]
Fig. 1.
Multiple sequence alignment of the putative
MAO segment responsible for substrate and inhibitor specificity.
The putative 161-amino acid segment responsible for specificity in
human MAO A and B (residues 215-375 in MAO A and 206-366 in MAO B) is
aligned from mouse, rat, and bovine MAO A, mouse and rat MAO B, and
trout MAO. The 15 amino acids that are nonconserved between MAO A and B
subtypes and concurrently conserved among all the different species
within a subtype are marked by asterisks. When trout MAO is
classified as an "MAO A subtype" because of similar kinetics (23),
the remaining five amino acids nonconserved between subtypes and
conserved among the different species of a subtype are
boxed. The three corresponding amino acid pairs selected for
reciprocal interchange by mutagenesis are indicated above the boxed
amino acids.
EXPERIMENTAL PROCEDURES
TOP
ABSTRACT
INTRODUCTION
EXPERIMENTAL PROCEDURES
RESULTS
DISCUSSION
REFERENCES
)-deprenyl were from Sigma.
5-[2-14C]hydroxytryptamine binoxalate (57.80 mCi/mmol)
and
-[ethyl-1-14C]phenylethylamine
hydrochloride (52.00 mCi/mmol) were from PerkinElmer Life
Sciences. Ro 41-1049 and Ro 16-6491 were from Research
Biochemicals International (Natick, MA). The pVL1392 transfer vector
and the linearized BaculoGoldTM DNA were from PharMingen (San Diego,
CA). Gentamycin, antibiotic-antimycotic solution, and fetal bovine serum were from Life Technologies, Inc. The 3,3'-diaminobenzidine was
from Pierce. The EagleSight software was from Stratagene.
80 °C.
4 to
10
11 M, and the IC50
values were determined by Hill analysis.
RESULTS
TOP
ABSTRACT
INTRODUCTION
EXPERIMENTAL PROCEDURES
RESULTS
DISCUSSION
REFERENCES
kcat and Km values of wild-type and mutant MAOs for the
substrates 5-HT and PEA
View larger version (24K):
[in a new window]
Fig. 2.
kcat/Km values of
wild-type MAO A and B and mutants MAO A-I335Y and MAO B-Y326I for the
substrates 5-HT and PEA. The specificity constants
(kcat/Km) for 5-HT and PEA
were calculated from the data in Table I. 5-HT is represented by the
black bar. PEA is represented by the gray
bar.
In contrast to MAO A, mutant MAO A-I335Y showed a higher kcat for PEA than for 5-HT. It also exhibited a 35-fold increase in its Km for 5-HT to 2801 µM (which is similar to the 5-HT Km of MAO B of 3891 µM); thus the Km for 5-HT was lower than for PEA (Table I). This produced a larger kcat/Km for PEA than for 5-HT (Fig. 2). In effect, A-I335Y acquired an MAO B-like substrate specificity.
Mutant B-Y326I had similar kcat values for 5-HT and PEA and, compared with MAO B, exhibited a 7-fold decrease in Km for 5-HT and a 5-fold increase in Km for PEA (Table I). The resulting kcat/Km values (Fig. 2) indicate that this mutant retained MAO B-like substrate specificities. However, the kcat/Km of the mutant was only about 75-fold higher for PEA than for 5-HT, compared with an ~40,000-fold difference in the MAO B wild type. Therefore even though B-Y326I retained a higher specificity for PEA than for 5-HT, it exhibited a significant shift in specificity toward MAO A. In summary, Ile-335 in MAO A and Tyr-326 in MAO B play an important role in the substrate specificity of human MAO A and B.
A switch in sensitivities for MAO A and B irreversible inhibitors
clorgyline and ()-deprenyl was also observed (Fig.
3). MAO A has an IC50 value
of 1.2 × 10
9 M for
clorgyline and 1.3 × 10
6 M
for deprenyl, whereas MAO B has an IC50 value of 6.3 × 10
7 M for clorgyline and
4.3 × 10
9 M for deprenyl.
Compared with MAO A, A-I335Y exhibited about a 6000-fold decrease in
sensitivity toward clorgyline (IC50 = 7.1 × 10
6 M) and a 10-fold increase in
sensitivity toward deprenyl (IC50 = 1.2 × 10
7 M) (Fig. 3). Thus the
inhibitor sensitivity of this MAO A mutant became MAO-B like.
Similarly, MAO B-Y326I was MAO A-like and was more sensitive to
clorgyline (IC50 = 2.8 × 10
8 M) than to deprenyl
(IC50 = 1.8 × 10
7
M) (Fig. 3). Therefore Ile-335 and Tyr-326 determine
clorgyline and deprenyl sensitivities.
|
We also studied enzyme sensitivity toward the reversible inhibitors Ro
41-1049 (MAO A-specific) and Ro 16-6491 (MAO B-specific). As shown in
Fig. 4A, A-I335Y had about a
4000-fold decreased sensitivity toward Ro 41-1049 compared with MAO A
(IC50 = 5.6 × 108
M for MAO A and 2.5 × 10
4
M for A-I335Y), becoming MAO B-like. Similarly, B-Y326I
exhibited a 1000-fold higher sensitivity for the MAO A-specific
inhibitor Ro 41-1049 compared with MAO B (IC50 = 2.5 × 10
4 M for MAO B and 2.5 × 10
6 M for B-Y326I) and became
more like MAO A. However, for the MAO B-specific inhibitor Ro
16-6491, both mutants exhibited a decrease in sensitivity when
compared with their parent enzyme, and no reversal in specificity was
observed.
|
The substrate and inhibitor specificities exhibited by A-I335Y and
B-Y326I suggest that specificity may be determined by the presence of
either an aliphatic or an aromatic side chain at this position. To
confirm this, we made mutants A-I335F and B-Y326V. A-I335F had
kcat/Km values of 3.2 × 104 for 5-HT and 0.03 for PEA, making it
similar to A-I335Y. In addition, B-Y326V had
kcat/Km values of 0.042 for
5-HT and 2.71 for PEA, which are similar to values for B-Y326I. For the
inhibitors, A-I335F had IC50 values of 6.8 ± 0.7 × 10
6 for clorgyline and 3.8 ± 0.4 × 10
7 for deprenyl, which are
similar to values for A-I335Y, and equivalently B-Y326V had
IC50 values of 2.4 ± 0.8 × 10
8 for clorgyline and 2.2 ± 0.5 × 10
7 for deprenyl, which are similar to
values for B-Y326I. These results show that it is the presence of
either an aromatic or aliphatic side chain that is responsible for the
inverse specificity effect.
The other two pairs of mutants, A-T245I and B-I236T and A-D328G and
B-G319D, showed a decrease in kcat values for
both 5-HT and PEA compared with their parent enzymes. A-T245I and
A-D328G showed a kcat of 12.6 ± 1.6 and
1.6 ± 0.2 min1, respectively, for 5-HT
(compared with 67.4 ± 3.4 min
1 for MAO
A wild type) and 4.2 ± 0.5 and 0.3 ± 0.1 min
1, respectively, for PEA (compared with
11.2 ± 0.4 min
1 for MAO A wild type).
B-I236T and B-G319D showed kcat values of
1.8 ± 0.1 and 2.2 ± 0.1 min
1,
respectively, for 5-HT (compared with 5.1 ± 0.1 for MAO B wild type) and 19.4 ± 0.8 and 14.3 ± 2.0 min
1, respectively, for PEA (compared with
98.4 ± 5.2 for MAO B wild type). However, they retained the
Km values for both substrates, thus resulting in
unaltered specificities.
Their IC50 values for clorgyline, deprenyl, Ro 49-1049, and Ro 16-6491 were also unchanged compared with their parent enzyme. These results suggest that the mutations decrease the catalytic activity by either directly or indirectly affecting the active site. However, they do not affect substrate or inhibitor specificity.
It should be noted that even though the substrate and inhibitor specificities of A-I335Y and B-Y326I were switched, their kinetic and inhibitory constants were not identical to those of the other MAO; the Km values for PEA were not greatly affected. The IC50 values for the inhibitors shift toward, but do not become identical to, the opposite MAO, and no specificity change was observed for Ro 16-6491. This suggests that other amino acids also play a role in determining substrate and inhibitor specificities.
In summary, the MAO A mutant A-I335Y acquired kinetic parameters
similar to those of MAO B. Similarly, the MAO B mutant B-Y326I acquired
kinetic parameters more like those of MAO A. Therefore our results
suggest that Ile-335 in MAO A and its corresponding residue in MAO B,
Tyr-326, play an important role in conferring substrate and inhibitor
preferences to human MAO A and B.
![]() |
DISCUSSION |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
Using site-directed mutagenesis, we constructed six MAO mutants by reciprocally interchanging three corresponding amino acid pairs in human MAO A and B within a region thought to be important for conferring substrate and inhibitor specificities. The corresponding mutant pair A-I335Y and B-Y326I exhibited opposite specificities compared with their parent enzymes, MAO A and B, respectively. A-I335Y became more like MAO B, whereas B-Y326I became more like MAO A. This suggests that Ile-335 in MAO A and Tyr-326 in MAO B have an important function in determining the specificities of human MAO A and B. Although the three-dimensional crystal structure of the enzyme is not available, we can explain the specificity switch effect in the following manner. It is possible that in human MAO A the binding of the substrate or inhibitor may be facilitated by direct hydrophobic interactions within the active site with Ile-335, whereas in human MAO B such binding may be mediated by aromatic stacking or hydrogen bonding with the hydroxyl group of Tyr-326. This view is supported by the observation that mutants A-I335F and B-Y326V also exhibited a change in specificity similar to the reciprocal mutants. It is also possible that these two amino acids influence the three-dimensional structure of the enzyme and affect substrate and inhibitor specificities.
However, this binding interaction may not be generalized to MAOs of other species. It was reported that switching Phe-208 and Ile-199 in rat MAO A and B, respectively, results in a partial inversion of specificities for some substrates and inhibitors (22). This result suggested that the structural feature responsible for determining specificity was the aromatic ring of Phe-208 in rat MAO A and the aliphatic side chain of Ile-199 in rat MAO B. This is the reverse of our present observation in human MAO, in which the aliphatic residue is in MAO A (Ile-335), and the aromatic residue is in MAO B (Tyr-326). Even though the data from the rat MAO mutants consisted only of Km values to ascertain substrate specificity and a single inhibitor concentration to determine inhibitor specificity, when compared with our results, these data may point to an MAO species difference in the way specificity is determined. In fact, we have previously found that the same amino acid substitutions made on human MAO, A-F208I and B-I199F, did not result in a change in specificities (21), as was observed in rats. Several reports indicate the existence of large differences in specificities among MAOs of the same subtype but from different mammalian species (27-30). Some oxadiazolone compounds have inhibitory potencies that vary four orders of magnitude between rat and bovine MAO B (28), whereas some antidepressant drugs show a B over A specificity in mouse and rat MAOs and the reverse specificity in rat and monkey MAOs (29).
These reports and our results suggest that the specificities of rat and human MAOs (and MAOs of other species) are determined by different amino acids. Indeed, a computer modeling study suggests spatial differences between the binding sites of MAO A and B and that one amino acid can be responsible for the binding of some but not all substrates and inhibitors (31). In addition, according to a three-dimensional MAO model3 based on the recently crystallized polyamine oxidase (32) that shares a 20% amino acid identity with MAO, Phe-208 and Ile-335 in MAO A and their equivalents in MAO B, Ile-199 and Tyr326, have adjacent side chains within a region close to the isoalloxazine moiety of the FAD. This suggests that the overall three-dimensional structure of human and rat MAOs may be similar but that specificity may be determined by different amino acids within the binding domain. MAO A and B may have similar amino acids involved in the active site; however, small variations in the three-dimensional structure result in different specificities.
The other four mutants (A-T245I, B-I236T, A-D328G, and B-G319D) did not result in any marked differences in their specificities when compared with the wild types. However their kcat values for both 5-HT and PEA were lower than those of the parent enzymes. Among them, a large decrease in kcat was observed in the two MAO A mutants, A-T245I and A-D328G. This indicates that these amino acid residues interact either directly or indirectly with the active site.
In summary, the present study identifies the corresponding amino acid
pair Ile-335 and Tyr-326 as critical for the substrate and inhibitor
specificities in human MAO A and B, respectively. Thr-245 and Asp-328
in MAO A and Ile-236 and Gly-319 in MAO A may interact with the active
site but do not determine specificity.
![]() |
FOOTNOTES |
---|
* This work was supported by National Institute of Mental Health Grants R01 MH37020, R37 MH39085 (MERIT Award), and Research Scientist Award K05 MH00796 and by the Welin Professorship.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: 1985 Zonal Ave., Los Angeles, CA 90089. Tel.: 323-442-1441; Fax: 323-442-3229; E-mail: jcshih@hsc.usc.edu.
Published, JBC Papers in Press, December 29, 2000, DOI 10.1074/jbc.M006972200
2 R. M. Geha, I. Rebrin, K. Chen, and J. C. Shih, unpublished data.
3 Johan Wouters, Belgium, personal communication.
![]() |
ABBREVIATIONS |
---|
The abbreviations used are:
MAO, monoamine
oxidase;
5-HT, 5-hydroxytryptamine;
PEA, -phenylethylamine..
![]() |
REFERENCES |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
1. |
Greenawalt, J. W.,
and Schnaitman, C.
(1970)
J. Cell Biol.
46,
173-179 |
2. | Nara, S., Igaue, I., Gomes, B., and Yasunobu, K. T. (1966) Biochem. Biophys. Res. Commun. 23, 324-328[Medline] [Order article via Infotrieve] |
3. | Shih, J. C., Chen, K., and Ridd, M. J. (1999) Annu. Rev. Neurosci. 22, 197-217[CrossRef][Medline] [Order article via Infotrieve] |
4. | Johnston, J. P. (1968) Biochem. Pharmacol. 17, 1285-1297[CrossRef][Medline] [Order article via Infotrieve] |
5. | Cesura, A. M., Bos, M., Galva, M. D., Imhof, R., and Da Prada, M. (1989) Mol. Pharmacol. 37, 358-366[Abstract] |
6. | Knoll, J., and Magyar, K. (1972) Adv. Biochem. Psychopharmacol. 5, 393-408[Medline] [Order article via Infotrieve] |
7. | Cesura, A. M., Imhof, R., Galva, M. D., Kettler, R., and Da Prada, M. (1988) Pharmacol. Res. Comm. 20, 51-61[Medline] [Order article via Infotrieve] |
8. | O' Carroll, A., Fowler, C. J., Phillips, J. P., Tobbia, I., and Tipton, K. F. (1983) Naunyn-Schmiedeberg's Arch. Pharmacol. 312, 51-55 |
9. | Bach, A. W. J., Lan, N. C., Johnson, D. L., Abell, C. W., Bembenck, M. E., Kwan, S. W., Seeburg, P. H., and Shih, J. C. (1988) Proc. Natl. Acad. Sci. U. S. A. 85, 4934-4938[Abstract] |
10. | Lan, N. C., Heinzmann, C., Gal, A., Klisak, I., Orth, U., Lai, E., Grimsby, J., Sparkes, R. S., Mohandas, T., and Shih, J. C. (1989) Genomics 4, 552-559[Medline] [Order article via Infotrieve] |
11. | Grimsby, J., Chen, K., Wang, L. J., Lan, N. C., and Shih, J. C. (1991) Proc. Natl. Acad. Sci. U. S. A. 88, 3637-3641[Abstract] |
12. | Brunner, H. G., Nelen, M., Breakfield, X. O., Ropers, H. H., and Van Oost, B. A. (1993) Science 262, 578-580[Medline] [Order article via Infotrieve] |
13. | Cases, O., Seif, I., Grimsby, J., Gaspar, P., Chen, K., Pournin, S., Müller, U., Aguet, M., Babinet, C., Shih, J. C., and De Maeyer, E. (1995) Science 268, 1763-1766[Medline] [Order article via Infotrieve] |
14. | Grimsby, J., Toth, M., Chen, K., Kumazawa, T., Klaidman, L., Adams, J. D., Karoum, F., Gal, J., and Shih, J. C. (1997) Nat. Genet. 17, 1-5[Medline] [Order article via Infotrieve] |
15. | Cesura, A. M., Gottowik, J., Lahm, H.-W., Lang, G., Imhof, R., Malherbe, P., Röthlisberger, U., and Da Prada, M. (1996) Eur. J. Biochem. 236, 996-1002[Abstract] |
16. | Chen, K., Wu, H.-F., and Shih, J. C. (1996) J. Neurochem. 66, 797-803[Medline] [Order article via Infotrieve] |
17. | Gottowik, J., Cesura, A. M., Malherbe, P., Lang, G., and Da Prada, M. (1993) FEBS Lett. 317, 152-156[CrossRef][Medline] [Order article via Infotrieve] |
18. | Gottowik, J., Malherbe, P., Jang, G., Da Prada, M., and Cesura, A. M. (1995) Eur. J. Biochem. 230, 934-942[Abstract] |
19. | Tsugeno, Y., Hirashiki, I., Ogata, F., and Ito, A. (1995) J. Biochem. (Tokyo) 118, 974-980[Abstract] |
20. | Wu, H.-F., Chen, K., and Shih, J. C. (1993) Mol. Pharmacol. 43, 888-893[Abstract] |
21. | Geha, R. M., Chen, K., and Shih, J. C. (2000) J. Neurochem. 75, 1304-1309[CrossRef][Medline] [Order article via Infotrieve] |
22. |
Tsugeno, Y.,
and Ito, A.
(1997)
J. Biol. Chem.
272,
14033-14036 |
23. | Chen, K., Wu, H.-F., Grimsby, J., and Shih, J. C. (1994) Mol. Pharmacol. 268, 1226-1233 |
24. | Crossen, R., and Gruenwald, S. (1996) Baculovirus Expression Vector Manual , 3rd Ed. , PharMingen, San Diego, CA |
25. | Schagger, H., and Jagow, G. (1987) Anal. Biochem. 166, 368-379[Medline] [Order article via Infotrieve] |
26. | Grimsby, J., Zentner, M., and Shih, J. C. (1996) Life Sci. 58, 777-787[CrossRef][Medline] [Order article via Infotrieve] |
27. | Glover, V., and Sandler, M. (1977) Nature 265, 80-81[Medline] [Order article via Infotrieve] |
28. | Krueger, M. J., Mazouz, F., Ramsay, R. R., Milcent, R., and Singer, T. P. (1995) Biochem. Biophys. Res. Commun. 206, 556-562[CrossRef][Medline] [Order article via Infotrieve] |
29. | Egashira, T., Takayama, F., and Yamanaka, Y. (1999) Jpn. J. Pharmacol. 81, 115-121[CrossRef][Medline] [Order article via Infotrieve] |
30. |
Inoue, H.,
Castagnoli, K.,
Van Der Schyf, C.,
Mabic, S.,
Igarashi, K.,
and Castagnoli, N.
(1999)
J. Pharmacol. Exp. Ther.
291,
856-864 |
31. | Veselovsky, A. V., Ivanov, A. S., and Medvedev, A. E. (1998) Biochemistry (Mosc.) 63, 1441-1446[Medline] [Order article via Infotrieve] |
32. | Binda, C., Coda, A., Angelini, R., Federico, R., Ascenzi, P., and Mattevi, A. (1999) Structure 7, 265-276[CrossRef][Medline] [Order article via Infotrieve] |