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Inhibition of fatty acid synthetases by the antibiotic cerulenin
BIOCHEMICAL
Vol. 48, No. 3, 1972
INHIBITION
AND BIOPHYSICAL RESEARCH COMMUNICATIONS
OF FATTY ACID SYNTHETASES ANTIBIOTIC CERULENIN
BY THE
D. Vance, I. Goldberg, 0. Mitsuhashi and K. Bloch Conant Laboratories, Harvard University Cambridge, Massachusetts 02138 U. S, A. and Satoshi bmura* and SetsuzS Kitasato Institute and Kitasato Tokyo, Japan Received
June
Summary:
The
inhibitor
26,1972 antibiotic
of fatty acid
cerulenin
is a potent
synthetase
systems
and from
rat liver.
Cerulenin
specifically
thioester
synthetase
(condensing
enzyme).
inhibition
of overall
fatty acid
Introduction: 0x0-6, fungi
Cerulenin,
and bacteria
(1,2)
(3).
lipid
molecules
interferes
with
the utilization
reduce
markedly cells
--in vivo
Addition
of the antibiotic
of 1-14C-acetate,
it was
suggested
beyond
the formation condensation
These
the growth
medium,
effects
may exert
common
of yeasts,
by the addition that cerulenin
to the biosynthesis
can also be demonstrated extracts
reduced
of
649
--in vitro
(5).
the incorporation into the fatty
On the basis
its inhibitory
and malonyl-CoA
to acetoacetyl-thioesters
0 I9 72 by Academic Press, Inc. in arty form reserved.
of a variety
it was proposed
by 60-900/o (5).
of acetyl-CoA
for the
(2S)(3R)2,3-epoxy-4-
can be overcome
effects interfering
of these
findings,
on some
step
most
likely
(5).
Present address: Visiting Research Professor, Wesleyan University, Middletown, Connecticut
of reproduction
may account
acyl
of this proposal, cerulenin was found to 14 of C-acetate into fatty acids and sterols
*
Copyright All rights
effect
microorganisms of P-keto
or 1, 3-14C-malonyl-CoA
fractions
that cerulenin
various
the activity
of the structure
(2-3 PM) to yeast
l-14C-acetyl-CoA
acid or non-saponifiable
their
(4).
This
of a precursor
the incorporation
of yeast
blocks
inhibition
In support
from
non-competitive
by the antibiotic.
to the growth
structures.
and apparently
isolated
inhibits
Since this
of certain
lipid
synthesis
an antibiotic
lo-dodecadienoylamide
several
Nomura University
Department of Chemistry, 06457 U. S. A.
with
Vol. 48, No. 3, 1972
BIOCHEMICAL
AND BIOPHYSICAL RESEARCH COMMUNICATIONS .*
Table I Percent Inhibition of Type I Fatty Acid Synthetases (Multienzyme Complexes) by Cerulenin
I
I
Euglena gracilis Yeast Type I
Corynebacterium diphtheriae
Mycobacterium phlei, Type1
4
13
90
100
82
8
14
94
100
88
56
20
51
97
93
79
97
99
40
I
I
100
87 95
We now report the effects of cerulenin on the activities of several crude or partially purified fatty acid synthetases from diverse sources,
In all but one
instance the incorporation of l4 C-malonyl-CoA into long chain fatty acids was strongly inhibited by low concentrations of the antibiotic (Tables I and II), Testing the effect of cerulenin on several of the partial reactions catalyzed by the high molecular weight synthetase of lVJ. m,
we have obtained evidence
that the antibiotic specifically inhibits the formation of P-ketoacyl thioesters, the reaction catalyzed by condensing enzyme. Experimental: Fatty Acid Synthetases - Fatty acid synthetase I from J& phlei ATCC356 was isolated as described (6), except that’the calcium phosphate step and sucrose gradient centrifugation were omitted and substituted by Bio-Gel A-5 chromatography.
g.
phlei synthetase II (palmityl CoA elongating system) was
prepared and assayed as described (7). Corynebacterium diphtheriae was kindly provided by Dr. A; Pappenheimer. This synthetase was isolated and purified by H. Knoche (8). Euglena gracilis strain Z was the source of three fatty acid synthetases.
650
BIOCHEMICAL
Vol. 48, No. 3, 1972
AND BIOPHYSICAL RESEARCH COMMUNICATIONS
Table
II
Percent Inhibition of ACP-Dependent (Type Fatty Acid Synthetases by Cerulenin
Mycobacterium
4
phlei
Euglena
0
8
95
20
99
II)
gracilis
99
77
100
87
0
93
40 100
10
200
10
*
palmityl-CoA
Enzyme
100
elongation
I is the multienzyme
synthetase
isolated
that catalyzes arachidate
from
green
Euglena
an ACP-dependent
and is present I&
complex
coli
Yeast
- The
both enzyme
and Enzyme cells
(9).
II the ACP-requiring Enzyme
elongation
of palmityl
in etiolated
and green
was fraction
- The synthetase
CoA to stearate Euglena
A as described
was partially
purified
III is an activity
cells
(14).
by Vagelos
as described
and
et al - -a
by Lynen
(10). (11).
Rat Liver - This enzyme was prepared according to Brady -et -0 al (12). -Fatty acid synthesis activity was assayed in the various systems under the conditions labelled
shown
III.
In all cases
2-14C-malonyl-CoA
was the
substrate. Condensing
described
in Table
by Kumar
enzyme
activity
-et -*al
(13).
of l& The
651
assay
phlei
synthetase
system
(total
I was assayed volume
as
of 0.2 ml)
pg
Protein,
*
All incubations
Palmityl-CoA
E. coli ACP, pM -
M
7.2,
pH
Kpo4 buffer,
(total vol.
l-2.5
0.1
0.5 ml) were
2-8
0.2
6
6
Dithiothreitol,
mM
2
19
carried
out for 20 min.
Q4-0.8
0.5
6
2
19
19
19
1
)IM
DPNK
125
25
100
2
at 37”
24-60
0.05
0.5
15
15
30
10
12-48
0.05
2
0. 5
15
15
10
0.05
2
0. 5
15
15
15
10
28-112
Euglena gracilis
For Fatty Acid Synthetases
Corynebacterium diphtheriae
Mixtures
25
100
Yeast
I
of Assay
FMN, pM
W
20
Malonyl CoA, pM
TPW
5
Acetyl CoA, @vl
Liver
Rat
Composition
Table III
2
0.1
5
1
30
30
20
300
5
2
74
0.2
-
200
200
30
100*
Mycobacterium pNei I II
BIOCHEMICAL
Vol. 48, No. 3, 1972
contained 1 mM
100 mM EDTA,
potassium
300 pM
bicarbonate
AND BIOPHYSICAL RESEARCH COMMUNICATIONS
phosphate
acetyl-CoA,
Q pC&nole).
buffer
(pH 7.0),
5 mM dithiothreitol, 14 and 100 mM C-sodium
50 pM malonyl-CoA,
The
reaction
was initiated
by addition
of 100 pg of
enzyme. Results: Cerulenin inhibits the synthesis of long-chain 14 C-malonyl-CoA incorporation --in vitro as effectively testing the antibiotic, we have chosen representatives synthetases
complexes
Saccharomyces
of these with
Fatty
of class elongating
systems.
inhibited phlei
from
I is also
chain
substrates
of
the synthetase
which
of g.
with
of yeast.
This
of fatty
in concentrations
malonyl-CoA-transacylase
are also
from
phlei
inhibits inhibition
from
Euglena
were
enzyme
of 100 ug per acyl
653
gracilis
(14) is
tested
The
I&
phlei it to
to the antibiotic.
with
model
to explore
was g.
activities
reductase.
synthetase
converting
sensitive
by Lynen
ml:
are
II is unexplained,
in order
source
following
as
ACP-dependent
of fatty acids,
of cerulenin
described
(10) and
of antibiotic.
is likewise
those
coli
In
to cerulenin
as the primer
enzyme
ACP,
ACP-dependent
synthesis
acid synthesis
of non-
of type II synthetases
synthetase
process
and p-keto
E.
even at 200 )l.g/ml
The
were
for added
non-aggregated
activity
and absence
The
Cerulenin
are as sensitive
palmityl-CoA
of the antibiotic.
acid
systems
the --de novo
(6).
These
and by the lack
(type II) consist
but the corresponding
catalyzing
(12).
In some instances
class
They
elongating
resistance
procedures
(ACP).
In a subgroup
activity
I and the assay
by antibiotic
(9).
systems
active
weights
(8))
I).
II).
in the presence
synthetase
degrees.
I (Table
reactions
of action
to varying
full
(C24) acids
Partial
protein
by cerulenin,
cerulenin
synthetase
carrier
gracilis
(7) retains
Apart
molecular
and show a requirement
The palmityl-CoA
exceptional
mode
For types
diphtheriae
(9) and rat liver
of the second
we have tested Euplena
gracilis
by high
(Table
enzymes
palmityl-CoA
longer
4 pg/ml
individual
the synthetases
I&
of the two major
(6), Corynebacterium
Euglena
systems
synthetases
photoauxotrophic
phlei
acyl
acid
category
from
(ll),
for external
the activities is complete
g.
are characterized
of a requirement
aggregated
from
cerevisiae
systems
strongly
as it does in viva.
by
the --de novo formation of long-chain fatty acids from and malonyl-CoA. Type I synthetases are represented by the
multienzyme
this
as judged
that catalyze
acetyl-CoA
enzyme
fatty acids
phlei
the
synthetase
(11) for the fatty were
not affected
acyl-CoA-transacylase, On the other
hand
BIOCHEMICAL
Vol. 48, No. 3, 1972
Q
I
I
AND BIOPHYSICAL RESEARCH COMMUNICATIONS
I
I
1
,
1.20--
-28
- 24 b
\f
l.OO.-
- 20
-E -
I 20
Figure 1.
I 40 jq/ml
I 60 Cerulenin
16
0 I 100
I 80
Effect of Cerulenin on Type I Fatty Acid Synthetase From IvJ. phlei. (o-o) fatty acid synthesis; (o-o) condensing enzyme activity.
condensing enzyme (,B-keto acyl thioester synthetase) was highly sensitive to 14 CO2 fixation into cerulenin (Fig. 1). This activity was assayed by measuring malonyl-CoA in the presence of acetyl-CoA
(13).
The cerulenin concentrations required for 50% inhibition of condensing enzyme activity
were 7 pg/ml as compared to 12 ug/ml for 50% inhibition of
overall fatty acid synthetase activity
(Fig, 1).
In one experiment, enzyme was
preincubated for 10 mm. with 10 pg/ml of cerulenin.
Fatty acid synthesis was
98% inhibited, whereas simultaneous incubation of inhibitor with the complete system gave only 57% inhibition.
Therefore,
inhibition appears to be non-
competitive. Discussion:
Cerulenin is a uniquely potent and apparently specific inhibitor of
fatty acid synthetases from all sources so far tested with the single exception noted.
Condensing enzyme appears to be the specific and possibly the only
target for the antibiotic.
Inhibition of this activity
is non-competitive suggesting
that cerulenin may combine non-covalently with an active enzyme site.
Cerulenin
affects fatty acid synthetases of all known types, whether they are multienzyme complexes or non-aggregated systems.
Moreover,
cerulenin interferes both
with -de novo fatty acid synthesis from acetyl-CoA and with chain elongation Chain-length specificity, therefore, does not appear starting with pslmityl-CoA.
654
BIOCHEMICAL
Vol. 48, No. 3, 1972
AND BIOPHYSICAL RESEARCH COMMUNICATIONS
to be a factor in the mode of cerulenin action.
Cerulenin should be a useful
tool for controlling the fatty acid content and composition of cells in studies on the function of lipids in membranes. far,
3-decynoyl-N-acetyl
The one agent used for this purpose so
cysteamine (15) interferes only with the biosynthesis of
unsaturated fatty acids in certain bacteria (16). Acknowledgments: This research was supported by grants-in-aid from the United States Public Health Service, the National Science Foundation, the Life Insurance Medical Research Fund, and the Eugene P. Higgins Trust Fund of Harvard University. Some of the experiments were carried out by Drs. H. Knoche, T. Esders and Mr. P. Flick. References:
1.
Gmura, S. , Katagiri, M. , Nakagawa, A., Sane, S., Nomura, S. and Hata, T. , J. Antibiotics, Ser. A 20, 349 (1967).
2.
Gmura, S., Nakagawa, A., Sekikawa, K, , Otani, M. and Hata, T. , Chem. Pharm. Bull., 11, 2361 (1969).
3.
Matsumae, A. , Nomura, S, and Hata, T, , J. Antibiotics, 1 (1964).
4.
Nomura, S., Horiuchi, T, , dmura, S. and Hata, T., in press.
5.
Nomura, S. , Horiuchi, T, , Hata, T. and dmura, S. , J. Antibiotics 2&, in press.
6.
Brindley,
7.
Matsumura, S., Brindley, D, N, and Bloch, K., Biochem. and Biophys. Res. Comm. 38, 369 (1970).
8.
Knoche, H. W. and Koths, K. E. , Fed. Proc. g,
9.
-10,
10.
Ser. A 11,
J. Biochem.,
D, H., Matsumura, S, and Bloch, K., Nature 224, 666 (1969).
1475 (1972).
J., Ernst-Fonberg, M. L. and Bloch, K., Arch. Biophys. 143, 384 (1971).
Biochem.
Vagelos, P. R, , Alberts, A. W. and Majerus, P. W., -Methods in Enzymology John F. Lowenstein, Editor, Vol XIV, p. 39 (Academic Press, 1969).
11.
Lynen, F., Methods in Enzymoloa, p. 17 (Acadesz, 1969).
12.
Brady, R. O., Bradley, R. M. and Trams, E, G, , J. Biol. Chem., 235, 3093 (1960).
655
John F. Lowenstein, Editor, Vol. XIV
Vol. 48, No. 3, 1972
BIOCHEMICAL
AND BIOPHYSICAL RESEARCH COMMUNICATIONS
13.
Kumar, S, , Dorsey, J. A., Muesing, R. A. and Porter, Biol. Chem, 245, 4732 (1970).
14.
Goldberg, I and Bloch, K., unpublished.
15.
Bloch, K,, The Enzymes, Vol. 5, 3rd - Ed., p. 441, P, Boyer, Editor, (Academic Press, 1971).
16.
Kass, L, , J. Biol. Chem., 243, 3223 (1968).
656
J. W., J.
Vol. 48, No. 3, 1972
INHIBITION
AND BIOPHYSICAL RESEARCH COMMUNICATIONS
OF FATTY ACID SYNTHETASES ANTIBIOTIC CERULENIN
BY THE
D. Vance, I. Goldberg, 0. Mitsuhashi and K. Bloch Conant Laboratories, Harvard University Cambridge, Massachusetts 02138 U. S, A. and Satoshi bmura* and SetsuzS Kitasato Institute and Kitasato Tokyo, Japan Received
June
Summary:
The
inhibitor
26,1972 antibiotic
of fatty acid
cerulenin
is a potent
synthetase
systems
and from
rat liver.
Cerulenin
specifically
thioester
synthetase
(condensing
enzyme).
inhibition
of overall
fatty acid
Introduction: 0x0-6, fungi
Cerulenin,
and bacteria
(1,2)
(3).
lipid
molecules
interferes
with
the utilization
reduce
markedly cells
--in vivo
Addition
of the antibiotic
of 1-14C-acetate,
it was
suggested
beyond
the formation condensation
These
the growth
medium,
effects
may exert
common
of yeasts,
by the addition that cerulenin
to the biosynthesis
can also be demonstrated extracts
reduced
of
649
--in vitro
(5).
the incorporation into the fatty
On the basis
its inhibitory
and malonyl-CoA
to acetoacetyl-thioesters
0 I9 72 by Academic Press, Inc. in arty form reserved.
of a variety
it was proposed
by 60-900/o (5).
of acetyl-CoA
for the
(2S)(3R)2,3-epoxy-4-
can be overcome
effects interfering
of these
findings,
on some
step
most
likely
(5).
Present address: Visiting Research Professor, Wesleyan University, Middletown, Connecticut
of reproduction
may account
acyl
of this proposal, cerulenin was found to 14 of C-acetate into fatty acids and sterols
*
Copyright All rights
effect
microorganisms of P-keto
or 1, 3-14C-malonyl-CoA
fractions
that cerulenin
various
the activity
of the structure
(2-3 PM) to yeast
l-14C-acetyl-CoA
acid or non-saponifiable
their
(4).
This
of a precursor
the incorporation
of yeast
blocks
inhibition
In support
from
non-competitive
by the antibiotic.
to the growth
structures.
and apparently
isolated
inhibits
Since this
of certain
lipid
synthesis
an antibiotic
lo-dodecadienoylamide
several
Nomura University
Department of Chemistry, 06457 U. S. A.
with
Vol. 48, No. 3, 1972
BIOCHEMICAL
AND BIOPHYSICAL RESEARCH COMMUNICATIONS .*
Table I Percent Inhibition of Type I Fatty Acid Synthetases (Multienzyme Complexes) by Cerulenin
I
I
Euglena gracilis Yeast Type I
Corynebacterium diphtheriae
Mycobacterium phlei, Type1
4
13
90
100
82
8
14
94
100
88
56
20
51
97
93
79
97
99
40
I
I
100
87 95
We now report the effects of cerulenin on the activities of several crude or partially purified fatty acid synthetases from diverse sources,
In all but one
instance the incorporation of l4 C-malonyl-CoA into long chain fatty acids was strongly inhibited by low concentrations of the antibiotic (Tables I and II), Testing the effect of cerulenin on several of the partial reactions catalyzed by the high molecular weight synthetase of lVJ. m,
we have obtained evidence
that the antibiotic specifically inhibits the formation of P-ketoacyl thioesters, the reaction catalyzed by condensing enzyme. Experimental: Fatty Acid Synthetases - Fatty acid synthetase I from J& phlei ATCC356 was isolated as described (6), except that’the calcium phosphate step and sucrose gradient centrifugation were omitted and substituted by Bio-Gel A-5 chromatography.
g.
phlei synthetase II (palmityl CoA elongating system) was
prepared and assayed as described (7). Corynebacterium diphtheriae was kindly provided by Dr. A; Pappenheimer. This synthetase was isolated and purified by H. Knoche (8). Euglena gracilis strain Z was the source of three fatty acid synthetases.
650
BIOCHEMICAL
Vol. 48, No. 3, 1972
AND BIOPHYSICAL RESEARCH COMMUNICATIONS
Table
II
Percent Inhibition of ACP-Dependent (Type Fatty Acid Synthetases by Cerulenin
Mycobacterium
4
phlei
Euglena
0
8
95
20
99
II)
gracilis
99
77
100
87
0
93
40 100
10
200
10
*
palmityl-CoA
Enzyme
100
elongation
I is the multienzyme
synthetase
isolated
that catalyzes arachidate
from
green
Euglena
an ACP-dependent
and is present I&
complex
coli
Yeast
- The
both enzyme
and Enzyme cells
(9).
II the ACP-requiring Enzyme
elongation
of palmityl
in etiolated
and green
was fraction
- The synthetase
CoA to stearate Euglena
A as described
was partially
purified
III is an activity
cells
(14).
by Vagelos
as described
and
et al - -a
by Lynen
(10). (11).
Rat Liver - This enzyme was prepared according to Brady -et -0 al (12). -Fatty acid synthesis activity was assayed in the various systems under the conditions labelled
shown
III.
In all cases
2-14C-malonyl-CoA
was the
substrate. Condensing
described
in Table
by Kumar
enzyme
activity
-et -*al
(13).
of l& The
651
assay
phlei
synthetase
system
(total
I was assayed volume
as
of 0.2 ml)
pg
Protein,
*
All incubations
Palmityl-CoA
E. coli ACP, pM -
M
7.2,
pH
Kpo4 buffer,
(total vol.
l-2.5
0.1
0.5 ml) were
2-8
0.2
6
6
Dithiothreitol,
mM
2
19
carried
out for 20 min.
Q4-0.8
0.5
6
2
19
19
19
1
)IM
DPNK
125
25
100
2
at 37”
24-60
0.05
0.5
15
15
30
10
12-48
0.05
2
0. 5
15
15
10
0.05
2
0. 5
15
15
15
10
28-112
Euglena gracilis
For Fatty Acid Synthetases
Corynebacterium diphtheriae
Mixtures
25
100
Yeast
I
of Assay
FMN, pM
W
20
Malonyl CoA, pM
TPW
5
Acetyl CoA, @vl
Liver
Rat
Composition
Table III
2
0.1
5
1
30
30
20
300
5
2
74
0.2
-
200
200
30
100*
Mycobacterium pNei I II
BIOCHEMICAL
Vol. 48, No. 3, 1972
contained 1 mM
100 mM EDTA,
potassium
300 pM
bicarbonate
AND BIOPHYSICAL RESEARCH COMMUNICATIONS
phosphate
acetyl-CoA,
Q pC&nole).
buffer
(pH 7.0),
5 mM dithiothreitol, 14 and 100 mM C-sodium
50 pM malonyl-CoA,
The
reaction
was initiated
by addition
of 100 pg of
enzyme. Results: Cerulenin inhibits the synthesis of long-chain 14 C-malonyl-CoA incorporation --in vitro as effectively testing the antibiotic, we have chosen representatives synthetases
complexes
Saccharomyces
of these with
Fatty
of class elongating
systems.
inhibited phlei
from
I is also
chain
substrates
of
the synthetase
which
of g.
with
of yeast.
This
of fatty
in concentrations
malonyl-CoA-transacylase
are also
from
phlei
inhibits inhibition
from
Euglena
were
enzyme
of 100 ug per acyl
653
gracilis
(14) is
tested
The
I&
phlei it to
to the antibiotic.
with
model
to explore
was g.
activities
reductase.
synthetase
converting
sensitive
by Lynen
ml:
are
II is unexplained,
in order
source
following
as
ACP-dependent
of fatty acids,
of cerulenin
described
(10) and
of antibiotic.
is likewise
those
coli
In
to cerulenin
as the primer
enzyme
ACP,
ACP-dependent
synthesis
acid synthesis
of non-
of type II synthetases
synthetase
process
and p-keto
E.
even at 200 )l.g/ml
The
were
for added
non-aggregated
activity
and absence
The
Cerulenin
are as sensitive
palmityl-CoA
of the antibiotic.
acid
systems
the --de novo
(6).
These
and by the lack
(type II) consist
but the corresponding
catalyzing
(12).
In some instances
class
They
elongating
resistance
procedures
(ACP).
In a subgroup
activity
I and the assay
by antibiotic
(9).
systems
active
weights
(8))
I).
II).
in the presence
synthetase
degrees.
I (Table
reactions
of action
to varying
full
(C24) acids
Partial
protein
by cerulenin,
cerulenin
synthetase
carrier
gracilis
(7) retains
Apart
molecular
and show a requirement
The palmityl-CoA
exceptional
mode
For types
diphtheriae
(9) and rat liver
of the second
we have tested Euplena
gracilis
by high
(Table
enzymes
palmityl-CoA
longer
4 pg/ml
individual
the synthetases
I&
of the two major
(6), Corynebacterium
Euglena
systems
synthetases
photoauxotrophic
phlei
acyl
acid
category
from
(ll),
for external
the activities is complete
g.
are characterized
of a requirement
aggregated
from
cerevisiae
systems
strongly
as it does in viva.
by
the --de novo formation of long-chain fatty acids from and malonyl-CoA. Type I synthetases are represented by the
multienzyme
this
as judged
that catalyze
acetyl-CoA
enzyme
fatty acids
phlei
the
synthetase
(11) for the fatty were
not affected
acyl-CoA-transacylase, On the other
hand
BIOCHEMICAL
Vol. 48, No. 3, 1972
Q
I
I
AND BIOPHYSICAL RESEARCH COMMUNICATIONS
I
I
1
,
1.20--
-28
- 24 b
\f
l.OO.-
- 20
-E -
I 20
Figure 1.
I 40 jq/ml
I 60 Cerulenin
16
0 I 100
I 80
Effect of Cerulenin on Type I Fatty Acid Synthetase From IvJ. phlei. (o-o) fatty acid synthesis; (o-o) condensing enzyme activity.
condensing enzyme (,B-keto acyl thioester synthetase) was highly sensitive to 14 CO2 fixation into cerulenin (Fig. 1). This activity was assayed by measuring malonyl-CoA in the presence of acetyl-CoA
(13).
The cerulenin concentrations required for 50% inhibition of condensing enzyme activity
were 7 pg/ml as compared to 12 ug/ml for 50% inhibition of
overall fatty acid synthetase activity
(Fig, 1).
In one experiment, enzyme was
preincubated for 10 mm. with 10 pg/ml of cerulenin.
Fatty acid synthesis was
98% inhibited, whereas simultaneous incubation of inhibitor with the complete system gave only 57% inhibition.
Therefore,
inhibition appears to be non-
competitive. Discussion:
Cerulenin is a uniquely potent and apparently specific inhibitor of
fatty acid synthetases from all sources so far tested with the single exception noted.
Condensing enzyme appears to be the specific and possibly the only
target for the antibiotic.
Inhibition of this activity
is non-competitive suggesting
that cerulenin may combine non-covalently with an active enzyme site.
Cerulenin
affects fatty acid synthetases of all known types, whether they are multienzyme complexes or non-aggregated systems.
Moreover,
cerulenin interferes both
with -de novo fatty acid synthesis from acetyl-CoA and with chain elongation Chain-length specificity, therefore, does not appear starting with pslmityl-CoA.
654
BIOCHEMICAL
Vol. 48, No. 3, 1972
AND BIOPHYSICAL RESEARCH COMMUNICATIONS
to be a factor in the mode of cerulenin action.
Cerulenin should be a useful
tool for controlling the fatty acid content and composition of cells in studies on the function of lipids in membranes. far,
3-decynoyl-N-acetyl
The one agent used for this purpose so
cysteamine (15) interferes only with the biosynthesis of
unsaturated fatty acids in certain bacteria (16). Acknowledgments: This research was supported by grants-in-aid from the United States Public Health Service, the National Science Foundation, the Life Insurance Medical Research Fund, and the Eugene P. Higgins Trust Fund of Harvard University. Some of the experiments were carried out by Drs. H. Knoche, T. Esders and Mr. P. Flick. References:
1.
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Gmura, S., Nakagawa, A., Sekikawa, K, , Otani, M. and Hata, T. , Chem. Pharm. Bull., 11, 2361 (1969).
3.
Matsumae, A. , Nomura, S, and Hata, T, , J. Antibiotics, 1 (1964).
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Nomura, S., Horiuchi, T, , dmura, S. and Hata, T., in press.
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Nomura, S. , Horiuchi, T, , Hata, T. and dmura, S. , J. Antibiotics 2&, in press.
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Brindley,
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Matsumura, S., Brindley, D, N, and Bloch, K., Biochem. and Biophys. Res. Comm. 38, 369 (1970).
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Knoche, H. W. and Koths, K. E. , Fed. Proc. g,
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D, H., Matsumura, S, and Bloch, K., Nature 224, 666 (1969).
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Biochem.
Vagelos, P. R, , Alberts, A. W. and Majerus, P. W., -Methods in Enzymology John F. Lowenstein, Editor, Vol XIV, p. 39 (Academic Press, 1969).
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Lynen, F., Methods in Enzymoloa, p. 17 (Acadesz, 1969).
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Brady, R. O., Bradley, R. M. and Trams, E, G, , J. Biol. Chem., 235, 3093 (1960).
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John F. Lowenstein, Editor, Vol. XIV
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Kumar, S, , Dorsey, J. A., Muesing, R. A. and Porter, Biol. Chem, 245, 4732 (1970).
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Goldberg, I and Bloch, K., unpublished.
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Bloch, K,, The Enzymes, Vol. 5, 3rd - Ed., p. 441, P, Boyer, Editor, (Academic Press, 1971).
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Kass, L, , J. Biol. Chem., 243, 3223 (1968).
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