第 24章 生物氧化
— 电子传递与氧化磷酸化作用
Electron Transport
and Oxidative Phosphorylation
一
分
解
代
谢
的
三
步
曲
二
线
粒
体
呼
吸
链
的
组
成
? 细胞内的线粒体
是生物氧化的主
要场所，主要功
能是将代谢物脱
下的氢通过多种
酶及辅酶所组成
的传递体系的传
递，最终与氧结
合生成水。
? 由供氢体、传递
体、受氢体以及
相应的酶催化系
统组成的这种代
谢途径一般称为
生物氧化还原链，
当受氢体是氧时，
称为呼吸链。
2,FADH2 呼吸链
SH
2
N A D H
N A D
F M N
F M N H
2 C o Q
C o Q H
2
C y t b C y t c 1 C y t c C y t a,a 3
O
2
H
2
O
Fe
2+
Fe
3+
N A D H 脱氢酶
Fe
3+
Fe
3+
Fe
2+
Fe
2+
Fe
2+
Fe
2+
2H
2+
2
1
O
-
脱氢酶
S
C o Q
C o Q H
2
C y t b C y t c 1 C y t c C y t a,a 3
O
2
H
2
O
Fe
2+
Fe
3+
Fe
3+
Fe
3+
Fe
2+
Fe
2+
Fe
2+
Fe
2+
2H
2+
2
1
O
-
F A D
F A D H
2
黄素脱氢酶
M
MH
2
1,NADH 呼吸链
三、呼吸链各组份在线粒体内膜上的定位
现代生物化学认为，整个呼吸链由四个蛋白复合组分和两个游离的
成分组成,
★ 四个蛋白复合组分,Complex Ⅰ, Complex Ⅱ, Complex Ⅲ,
Complex Ⅳ
★ 两个游离成分,辅酶 Q和细胞色素 C
? An estimate of 10 protons are pumped for oxidizing one
NADH and 6 for one FADH2 accompanying the electron
flow through complexes I,III and IV,
? Conformational changes induced by electron transferring is
believed to be coupled to proton pumping (however,the
actual mechanisms is yet revealed!),
? In actively respiring mitochondria,the measured ?pH is
about 0.75 and difference in electrical potential (??) is
about 0.15-0.2 V,
? The energy stored in such an H+ gradient can be
used to synthesize ATP or to do other work,
四,呼吸链的电子传递在线粒体内膜两侧形成质
子梯度 （ A proton gradient across the inner membrane of
mitochondria is generated using the electron motive force ）
? Measurement of the standard reduction potential (?E`0)),
Electrons tend to transfer from low ?E`0 carriers to high ?E`0 carriers
(but may deviate from this in real cells),
? Oxidation kinetics studies,Full reduction followed by sudden O2
introduction; earlier oxidation,closer to the end of the respiratory
chain; using rapid and sensitive spectrophotometric techniques to
follow the oxidation of the cytochromes,which have different
wavelength of maximal absorption,
? Effects of various specific inhibitors,those before the blocked step
should be reduced and those after be oxidized,
? Isolation and characterization of each of the multiprotein
complexes,specific electron donors and acceptors can be determined
for portions of the chain,
五、呼吸链顺序的实验证据 （ The order of the many
electron carriers on the respiratory chain have been
elucidated via various studies）
六
线
粒
体
中
呼
吸
链
电
子
传
递
与AT
P
合
成
的
偶
联
? ATP would not be synthesized when
only ADP and Pi are added in isolated
mitochondria suspensions,
? O2 consumption,an indication of
electron flow,was detected when a
reductant (e.g.,succinate) is added,
accompanied by an increase of ATP
synthesis,
? Both O2 consumption and ATP
synthesis were suppressed when
inhibitors of respiratory chain (e.g.,
cyanide,CO,or antimycin A) was
added,
? ATP synthesis depends on the occurrence of
electron flow in mitochondria,
? O2 consumption was neither observed if ADP
was not added to the suspension,although a
reductant is provided,
? The O2 consumption was also not observed in
the presence of inhibitors of ATP synthase
(e.g.,oligomycin or venturicidin),
? Electron flow also depends on ATP synthesis!
七、电子传递与 ATP合成的偶联机制 （ The chemiosmotic
model was proposed to explain the coupling of electron flow and
ATP synthesis）
?★ First proposed in 1961 by Peter Michell (a British),
?★ Energy released from electron transferring is hypothesized
to be first used to pump protons from the mitochondrial matrix
to the intermembrane space (or from stroma to thylakoid
lumen in chloroplasts),thus generating a proton gradient
across the inner membrane; such a proton-motive force then
drives ATP synthesis by moving protons back into the matrix
via the ATP synthase-- The chemiosmotic model,
?★ The model was initially opposed by virtually all
researchers working in oxidative phosphorylation and
photosynthesis,
八
化
学
渗
透
学
说
的
实
验
证
据
? A closed membrane system is essential
for ATP synthesis but not for the
electron flow (tested with detergent or
physical shearing,
? Hydrophobic weak acids (DNP and
FCCP) and ionophores (valinomycin)
were found to be able to uncouple ATP
synthesis from electron transferring,
? The transmembrane proton pumping
has been experimentally detected,pH
in the intermembrane space was found to
decrease when electron flow occurs
(more protons are pumped when NADH,
rather than succinate,is utilized as
reductant),
? An artificially imposed electrochemical gradient across
the chloroplast thylakoid membrane and inner
mitochondrial membrane alone (both were performed using
sub-organelle vesicles) were found to drive ATP synthesis
(with the ATP synthase present),
? The across-membrane proton gradient was thus finally
accepted as the driving force for ATP synthesis,the
chemiosmotic model was accepted as a theory!
? The chemiosmotic theory unified the apparently disparate
energy transduction processes as oxidative phosphorylation,
photophosphorylation,active transport across membrane
and the motion of bacterial flagella,
? The F1 part consists of nine subunits of five types,
a3b3gde,
? The knoblike F1 portion is a hexamer of
alternating a and b subunits (arranged like the
segments of an orange),which sits atop the
single rod-shaped g subunit,
? The Fo portion consists three types of subunits,
ab2c10-12,
? The c subunits,each forming two transmembrane
helices,form a donut-shaped ring in the plane of
the membrane,
ATP synthase comprises a proton channel
(Fo) and a ATPase (F1)
? The leg-and-foot-shaped ge subunits stands
firmly on the ring of c subunits,
? The two b subunits of Fo seem to connect to the
ab hexamer via the d subunit of F1,
? The proton channel is believed to lie between the
a subunit and the ring of c subunits,
? X-ray crystallography revealed that the three b
subunits of F1 assumes three different
conformations,with bound ADP,ATP analog,or
empty respectively (John Walker,1994,Nature,
370:621-628)!
The binding-change model was proposed to explain
the action mechanism of ATP synthase
? The model was proposed by Paul Boyer in 1973
(PNAS,70:2837-2839),based on kinetic and binding
studies (before the 3-D structure of bovine F1 or
yeast FoF1 was determined),
? Downhill proton movement through Fo will drive the
rotation of the c-subunit ring and the asymmetrical
g subunits,which will cause each of the three b
subunits to interconvert between the three
conformations,as a result,each of them take turns to
take up ADP + Pi,synthesize ATP,and release ATP,
? Rotations of the g subunit and the c subunits of the
F1 unit in three discrete steps of 120o (powered by
ATP hydrolysis catalyzed by the b subunits) have
been directly observed using fluorescence
microscopy by Dr,Kazuhiko Kinosita in 1997
(Nature,386:299-302),
? The estimation of H+ consumption for each ATP
formed is 4 (among which one is consumed for Pi
transport),thus about 2.5 ATP/NADH,1.5
ATP/FADH2,
? The chemiosmotic coupling allows nonintegral
stoichiometries of O2 consumption (or NADH and
FADH2 oxidation) and ATP synthesis,
NADH Dehydrogenase
(complex I)
? Also named as NADH:ubiquinone oxidoreductase or
NADH-Q reductase,
? A,L” shaped 850 kD multimeric protein complex of 42
different subunits (larger than a ribosome!),
? Polypeptides encoded by both genomes,
? FMN,Fe-S centers act as prosthetic groups,
? Exergonic electron transferring is coupled to endergonic
proton pumping ( with 4 H+ pumped from the matrix
side to intermembrane space per electron pair
transferred),with mechanism unknown,
? Final electron acceptor is ubiquinone (coenzyme Q),
NADH 中的电子通过 NADH
dehydrogenase (complex I)进入呼吸链
Ubiquinone (Q)
accepts electrons
from both NADH
and FADH2 in the
respiratory chain
?Fat soluble
benzoquinone with a very
long isoprenoid side
chain; can accept one or
two electrons,forming
radical semiquinone or
ubiquinol (QH2); QH2
diffuses to the next
complex (III); the only
electron carrier not bound
to a protein,
CoQ是一个脂溶性
可移动的电子／质子
载体
Cytochrome bc1 complex (complex III)
Electron path in complex III
? Also called cytochrome c reductase or cytochrome bc1
complex,
? A 250 kD multiprotein complex of 11 subunits,
? Complete 3-D structure was determined in 1997!
? The functional core consists of three subunits,
cytochrome b (with two hemes,bH and bL); an Fe-S
protein; and cytochrome c1 (with the heme group
covalently bound to protein via two thioether bonds),
? Two-electron carrier QH2 passes one electron to the one-
electron carrier Fe-S center,then to the heme C group in
Cyt c1,and finally to the heme C group of Cyt c; the
other electron to bL,bH,and finally to an Q or Q.- via a
so-called,Q cycle”,
? Cytochrome c,a soluble protein located in the
intermembrane space,will move to complex IV,
细胞色素 c（ cyt.c）
它是电子传递链中一
个独立的蛋白质电子
载体, 位于线粒体内
膜外表, 属于膜周蛋
白, 易溶于水 。 它与
细胞色素 c1含有相同
的辅基, 但是蛋白组
成则有所不同 。 在电
子传递过程中, cyt,
c通过 Fe3+ ? Fe2+ 的
互变起电子传递中间
体作用 。
? Electrons of
Cyt c are
transferred to
O2 on
cytochrome
oxidase
(complex IV)
? A 204 kD 13-subunit protein complex,with structure
determined in 1996,
? Three subunits are probably critical to the function,
? Three copper ions (2 CuA,1CuB),two heme A groups (a
and a3) act as electron carriers in complex IV,
? Four electrons need to be transferred to reduce one O2
molecule at the,Fe-CuB center” (via peroxy intermediates)
of complex IV to form 2 H2O,
?Four,substrate” protons are consumed from the N
side for every four electrons transferred to one O2
molecule,
? One proton is pumped out from the N to P side for
each electron to be transferred by an yet defined
mechanism,
The three critical subunits of
cytochrome oxidase (complex IV),
2CuA
CuB
Heme a
Heme a3 CuA
CuA
Electron carriers may have an order of increasing E`0
Various inhibitors generate various
patterns of reduced/oxidized carriers
Reduced Oxidized
Reduced Oxidized
Reduced
? Resolution of functional complexes of the
respiratory chain,The outer mitochondrial
membrane is first removed by treatment
with the detergent digitonin,Fragments of
inner membrane are then obtained by
osmotic rupture of the mitochondria,and
the fragments are gently dissolved in a
second detergent,The resulting mixture of
inner membrane proteins is resolved by
ion-exchange chromatography into
different complexes (I through IV) of the
respiratory chain,each with its unique
protein composition,and the enzyme ATP
synthase (sometimes called Complex V),
The isolated Complexes I through IV
catalyze transfers between donors (NADH
and succinate),intermediate carriers (UQ
and cytochrome c),and O2,as shown,
ATP synthase,which is composed of
peripheral (F1) and integral (F0) proteins
(discussed later in this chapter),has only
ATP-hydrolyzing ;ATPase),not ATP-
synthesizing,activity in vitro,
Electron transfer was found to be
obligatorily coupled to ATP Synthesis
in isolated mitochondria suspensions,
neither occurs without the other,
The chemiosmotic
model by Mitchell
ATP synthesized
DNP,a hydrophobic weak acid,uncouples
ATP synthesis from electron flow
DNP and CCCP
are able to
dissipate the
proton gradient
The artificially
imposed proton
gradient alone
was found to drive
ATP synthesis!
1,破碎线粒体内膜，获取
反转的内膜脂质体，含
有呼吸链和 ATP合成酶。
2,用胰蛋白酶或尿素处理，
使 F1部分从线粒体内膜
上解离，并从此体系中
分离除去，此时，只传
递电子不合成 ATP,
3,重新加入 F1,使之与 F0
结合，传递电子的同时
合成 ATP,
The ATP synthase
comprises a proton
channel (Fo) and a
ATPase (F1)
The ten c subunits of Fo
The g subunit of F1
a
b
(The yeast FoF1 structure)
Rod-shaped g subunit,
ADP App(NH)p
Empty
Each b sununit of ATP synthase can
assume three different conformations!
b-ATP
b-ADP b-empty
The binding-change
model proposed by
Paul Boyer
g
g g
Rotation of the g
subunit and the
ring of c subunits
in the FoF1 complex
was observed by
in vitro studies
using fluorescence
microscopy
Rotation of the g subunit and the ring of c subunits
in the FoF1 complex was directly observed by in vitro
studies using fluorescence microscopy
— 电子传递与氧化磷酸化作用
Electron Transport
and Oxidative Phosphorylation
一
分
解
代
谢
的
三
步
曲
二
线
粒
体
呼
吸
链
的
组
成
? 细胞内的线粒体
是生物氧化的主
要场所，主要功
能是将代谢物脱
下的氢通过多种
酶及辅酶所组成
的传递体系的传
递，最终与氧结
合生成水。
? 由供氢体、传递
体、受氢体以及
相应的酶催化系
统组成的这种代
谢途径一般称为
生物氧化还原链，
当受氢体是氧时，
称为呼吸链。
2,FADH2 呼吸链
SH
2
N A D H
N A D
F M N
F M N H
2 C o Q
C o Q H
2
C y t b C y t c 1 C y t c C y t a,a 3
O
2
H
2
O
Fe
2+
Fe
3+
N A D H 脱氢酶
Fe
3+
Fe
3+
Fe
2+
Fe
2+
Fe
2+
Fe
2+
2H
2+
2
1
O
-
脱氢酶
S
C o Q
C o Q H
2
C y t b C y t c 1 C y t c C y t a,a 3
O
2
H
2
O
Fe
2+
Fe
3+
Fe
3+
Fe
3+
Fe
2+
Fe
2+
Fe
2+
Fe
2+
2H
2+
2
1
O
-
F A D
F A D H
2
黄素脱氢酶
M
MH
2
1,NADH 呼吸链
三、呼吸链各组份在线粒体内膜上的定位
现代生物化学认为，整个呼吸链由四个蛋白复合组分和两个游离的
成分组成,
★ 四个蛋白复合组分,Complex Ⅰ, Complex Ⅱ, Complex Ⅲ,
Complex Ⅳ
★ 两个游离成分,辅酶 Q和细胞色素 C
? An estimate of 10 protons are pumped for oxidizing one
NADH and 6 for one FADH2 accompanying the electron
flow through complexes I,III and IV,
? Conformational changes induced by electron transferring is
believed to be coupled to proton pumping (however,the
actual mechanisms is yet revealed!),
? In actively respiring mitochondria,the measured ?pH is
about 0.75 and difference in electrical potential (??) is
about 0.15-0.2 V,
? The energy stored in such an H+ gradient can be
used to synthesize ATP or to do other work,
四,呼吸链的电子传递在线粒体内膜两侧形成质
子梯度 （ A proton gradient across the inner membrane of
mitochondria is generated using the electron motive force ）
? Measurement of the standard reduction potential (?E`0)),
Electrons tend to transfer from low ?E`0 carriers to high ?E`0 carriers
(but may deviate from this in real cells),
? Oxidation kinetics studies,Full reduction followed by sudden O2
introduction; earlier oxidation,closer to the end of the respiratory
chain; using rapid and sensitive spectrophotometric techniques to
follow the oxidation of the cytochromes,which have different
wavelength of maximal absorption,
? Effects of various specific inhibitors,those before the blocked step
should be reduced and those after be oxidized,
? Isolation and characterization of each of the multiprotein
complexes,specific electron donors and acceptors can be determined
for portions of the chain,
五、呼吸链顺序的实验证据 （ The order of the many
electron carriers on the respiratory chain have been
elucidated via various studies）
六
线
粒
体
中
呼
吸
链
电
子
传
递
与AT
P
合
成
的
偶
联
? ATP would not be synthesized when
only ADP and Pi are added in isolated
mitochondria suspensions,
? O2 consumption,an indication of
electron flow,was detected when a
reductant (e.g.,succinate) is added,
accompanied by an increase of ATP
synthesis,
? Both O2 consumption and ATP
synthesis were suppressed when
inhibitors of respiratory chain (e.g.,
cyanide,CO,or antimycin A) was
added,
? ATP synthesis depends on the occurrence of
electron flow in mitochondria,
? O2 consumption was neither observed if ADP
was not added to the suspension,although a
reductant is provided,
? The O2 consumption was also not observed in
the presence of inhibitors of ATP synthase
(e.g.,oligomycin or venturicidin),
? Electron flow also depends on ATP synthesis!
七、电子传递与 ATP合成的偶联机制 （ The chemiosmotic
model was proposed to explain the coupling of electron flow and
ATP synthesis）
?★ First proposed in 1961 by Peter Michell (a British),
?★ Energy released from electron transferring is hypothesized
to be first used to pump protons from the mitochondrial matrix
to the intermembrane space (or from stroma to thylakoid
lumen in chloroplasts),thus generating a proton gradient
across the inner membrane; such a proton-motive force then
drives ATP synthesis by moving protons back into the matrix
via the ATP synthase-- The chemiosmotic model,
?★ The model was initially opposed by virtually all
researchers working in oxidative phosphorylation and
photosynthesis,
八
化
学
渗
透
学
说
的
实
验
证
据
? A closed membrane system is essential
for ATP synthesis but not for the
electron flow (tested with detergent or
physical shearing,
? Hydrophobic weak acids (DNP and
FCCP) and ionophores (valinomycin)
were found to be able to uncouple ATP
synthesis from electron transferring,
? The transmembrane proton pumping
has been experimentally detected,pH
in the intermembrane space was found to
decrease when electron flow occurs
(more protons are pumped when NADH,
rather than succinate,is utilized as
reductant),
? An artificially imposed electrochemical gradient across
the chloroplast thylakoid membrane and inner
mitochondrial membrane alone (both were performed using
sub-organelle vesicles) were found to drive ATP synthesis
(with the ATP synthase present),
? The across-membrane proton gradient was thus finally
accepted as the driving force for ATP synthesis,the
chemiosmotic model was accepted as a theory!
? The chemiosmotic theory unified the apparently disparate
energy transduction processes as oxidative phosphorylation,
photophosphorylation,active transport across membrane
and the motion of bacterial flagella,
? The F1 part consists of nine subunits of five types,
a3b3gde,
? The knoblike F1 portion is a hexamer of
alternating a and b subunits (arranged like the
segments of an orange),which sits atop the
single rod-shaped g subunit,
? The Fo portion consists three types of subunits,
ab2c10-12,
? The c subunits,each forming two transmembrane
helices,form a donut-shaped ring in the plane of
the membrane,
ATP synthase comprises a proton channel
(Fo) and a ATPase (F1)
? The leg-and-foot-shaped ge subunits stands
firmly on the ring of c subunits,
? The two b subunits of Fo seem to connect to the
ab hexamer via the d subunit of F1,
? The proton channel is believed to lie between the
a subunit and the ring of c subunits,
? X-ray crystallography revealed that the three b
subunits of F1 assumes three different
conformations,with bound ADP,ATP analog,or
empty respectively (John Walker,1994,Nature,
370:621-628)!
The binding-change model was proposed to explain
the action mechanism of ATP synthase
? The model was proposed by Paul Boyer in 1973
(PNAS,70:2837-2839),based on kinetic and binding
studies (before the 3-D structure of bovine F1 or
yeast FoF1 was determined),
? Downhill proton movement through Fo will drive the
rotation of the c-subunit ring and the asymmetrical
g subunits,which will cause each of the three b
subunits to interconvert between the three
conformations,as a result,each of them take turns to
take up ADP + Pi,synthesize ATP,and release ATP,
? Rotations of the g subunit and the c subunits of the
F1 unit in three discrete steps of 120o (powered by
ATP hydrolysis catalyzed by the b subunits) have
been directly observed using fluorescence
microscopy by Dr,Kazuhiko Kinosita in 1997
(Nature,386:299-302),
? The estimation of H+ consumption for each ATP
formed is 4 (among which one is consumed for Pi
transport),thus about 2.5 ATP/NADH,1.5
ATP/FADH2,
? The chemiosmotic coupling allows nonintegral
stoichiometries of O2 consumption (or NADH and
FADH2 oxidation) and ATP synthesis,
NADH Dehydrogenase
(complex I)
? Also named as NADH:ubiquinone oxidoreductase or
NADH-Q reductase,
? A,L” shaped 850 kD multimeric protein complex of 42
different subunits (larger than a ribosome!),
? Polypeptides encoded by both genomes,
? FMN,Fe-S centers act as prosthetic groups,
? Exergonic electron transferring is coupled to endergonic
proton pumping ( with 4 H+ pumped from the matrix
side to intermembrane space per electron pair
transferred),with mechanism unknown,
? Final electron acceptor is ubiquinone (coenzyme Q),
NADH 中的电子通过 NADH
dehydrogenase (complex I)进入呼吸链
Ubiquinone (Q)
accepts electrons
from both NADH
and FADH2 in the
respiratory chain
?Fat soluble
benzoquinone with a very
long isoprenoid side
chain; can accept one or
two electrons,forming
radical semiquinone or
ubiquinol (QH2); QH2
diffuses to the next
complex (III); the only
electron carrier not bound
to a protein,
CoQ是一个脂溶性
可移动的电子／质子
载体
Cytochrome bc1 complex (complex III)
Electron path in complex III
? Also called cytochrome c reductase or cytochrome bc1
complex,
? A 250 kD multiprotein complex of 11 subunits,
? Complete 3-D structure was determined in 1997!
? The functional core consists of three subunits,
cytochrome b (with two hemes,bH and bL); an Fe-S
protein; and cytochrome c1 (with the heme group
covalently bound to protein via two thioether bonds),
? Two-electron carrier QH2 passes one electron to the one-
electron carrier Fe-S center,then to the heme C group in
Cyt c1,and finally to the heme C group of Cyt c; the
other electron to bL,bH,and finally to an Q or Q.- via a
so-called,Q cycle”,
? Cytochrome c,a soluble protein located in the
intermembrane space,will move to complex IV,
细胞色素 c（ cyt.c）
它是电子传递链中一
个独立的蛋白质电子
载体, 位于线粒体内
膜外表, 属于膜周蛋
白, 易溶于水 。 它与
细胞色素 c1含有相同
的辅基, 但是蛋白组
成则有所不同 。 在电
子传递过程中, cyt,
c通过 Fe3+ ? Fe2+ 的
互变起电子传递中间
体作用 。
? Electrons of
Cyt c are
transferred to
O2 on
cytochrome
oxidase
(complex IV)
? A 204 kD 13-subunit protein complex,with structure
determined in 1996,
? Three subunits are probably critical to the function,
? Three copper ions (2 CuA,1CuB),two heme A groups (a
and a3) act as electron carriers in complex IV,
? Four electrons need to be transferred to reduce one O2
molecule at the,Fe-CuB center” (via peroxy intermediates)
of complex IV to form 2 H2O,
?Four,substrate” protons are consumed from the N
side for every four electrons transferred to one O2
molecule,
? One proton is pumped out from the N to P side for
each electron to be transferred by an yet defined
mechanism,
The three critical subunits of
cytochrome oxidase (complex IV),
2CuA
CuB
Heme a
Heme a3 CuA
CuA
Electron carriers may have an order of increasing E`0
Various inhibitors generate various
patterns of reduced/oxidized carriers
Reduced Oxidized
Reduced Oxidized
Reduced
? Resolution of functional complexes of the
respiratory chain,The outer mitochondrial
membrane is first removed by treatment
with the detergent digitonin,Fragments of
inner membrane are then obtained by
osmotic rupture of the mitochondria,and
the fragments are gently dissolved in a
second detergent,The resulting mixture of
inner membrane proteins is resolved by
ion-exchange chromatography into
different complexes (I through IV) of the
respiratory chain,each with its unique
protein composition,and the enzyme ATP
synthase (sometimes called Complex V),
The isolated Complexes I through IV
catalyze transfers between donors (NADH
and succinate),intermediate carriers (UQ
and cytochrome c),and O2,as shown,
ATP synthase,which is composed of
peripheral (F1) and integral (F0) proteins
(discussed later in this chapter),has only
ATP-hydrolyzing ;ATPase),not ATP-
synthesizing,activity in vitro,
Electron transfer was found to be
obligatorily coupled to ATP Synthesis
in isolated mitochondria suspensions,
neither occurs without the other,
The chemiosmotic
model by Mitchell
ATP synthesized
DNP,a hydrophobic weak acid,uncouples
ATP synthesis from electron flow
DNP and CCCP
are able to
dissipate the
proton gradient
The artificially
imposed proton
gradient alone
was found to drive
ATP synthesis!
1,破碎线粒体内膜，获取
反转的内膜脂质体，含
有呼吸链和 ATP合成酶。
2,用胰蛋白酶或尿素处理，
使 F1部分从线粒体内膜
上解离，并从此体系中
分离除去，此时，只传
递电子不合成 ATP,
3,重新加入 F1,使之与 F0
结合，传递电子的同时
合成 ATP,
The ATP synthase
comprises a proton
channel (Fo) and a
ATPase (F1)
The ten c subunits of Fo
The g subunit of F1
a
b
(The yeast FoF1 structure)
Rod-shaped g subunit,
ADP App(NH)p
Empty
Each b sununit of ATP synthase can
assume three different conformations!
b-ATP
b-ADP b-empty
The binding-change
model proposed by
Paul Boyer
g
g g
Rotation of the g
subunit and the
ring of c subunits
in the FoF1 complex
was observed by
in vitro studies
using fluorescence
microscopy
Rotation of the g subunit and the ring of c subunits
in the FoF1 complex was directly observed by in vitro
studies using fluorescence microscopy
