Title: Oxidative phosphorylation
1Oxidative Phosphorylation
R. C. Gupta Professor and Head Dept. of
Biochemistry National Institute of Medical
Sciences Jaipur, India
2Energy
E M B - R C G
EMB-RCG
3- The ultimate source of energy is the food that we
consume - Carbohydrates, lipids and proteins present in
food provide us energy - These are present in food in the form of
large complex molecules
4E M B - R C G
EMB-RCG
5(No Transcript)
6(No Transcript)
7High-energy phosphates
E M B - R C G
There must be some way of storing energy so that
it may be readily available when needed
The energy released during catabolism is captured
in the form of high-energy phosphates
8E M B - R C G
9 E M B - R C G
10 E M B - R C G
11Adenosine triphosphate
12(No Transcript)
13Standard free energy (DGo) of hydrolysis of some
important organic phosphates (kcal/mol)
These are the values of DGo obtained in standard
laboratory conditions of 1M reactant
concentration at pH 7.0 at 25C values obtained
in living cells (DGo) are different as the
reactant concentrations, pH and temperature
are different
14 E M B - R C G
15 E M B - R C G
EMB-RCG
16 E M B - R C G
17E M B - R C G
EMB-RCG
18 E M B - R C G
19 Oxidation and reduction
E M B - R C G
20 E M B - R C G
21(No Transcript)
22 E M B - R C G
23 E M B - R C G
24 E M B - R C G
25E M B - R C G
EMB-RCG
26(No Transcript)
27 Redox potential
E M B - R C G
28 E M B - R C G
29 Measurement of redox potential
E M B - R C G
30 E M B - R C G
31(No Transcript)
32 E M B - R C G
33(No Transcript)
34 Enzymes involved in biological oxidation
E M B - R C G
35 Oxidases
Oxidases transfer hydrogen from a substrate to
oxygen forming water or hydrogen peroxide
E M B - R C G
The enzymes forming water are metallo-enzymes
that usually contain copper e.g. cytochrome
oxidase and tyrosinase
The general reaction catalysed by oxidases is
AH2 ½ O2 ? A H2O
36 E M B - R C G
37 Dehydrogenases
E M B - R C G
38(No Transcript)
39Prosthetic group Examples
NAD Lactate dehydrogenase, malate dehydrogenase etc
NADP Glucose-6-phosphate dehydro-genase, 6-phosphogluconate dehydrogenase etc
Flavin nucleotides Succinate dehydrogenase, acyl CoA dehydrogenase etc
Iron- porphyrin Cytochrome a, cytochrome b etc
E M B - R C G
40 Hydroperoxidases
E M B - R C G
41(No Transcript)
42E M B - R C G
43 Mono-oxygenases
E M B - R C G
44 E M B - R C G
45(No Transcript)
46E M B - R C G
EMB-RCG
47E M B - R C G
EMB-RCG
48(No Transcript)
49(No Transcript)
50(No Transcript)
51Mitochondral hydroxylase system
E M B - R C G
52E M B - R C G
53Adrenodoxin possesses an iron-sulphur centre
having catalytic activity
E M B - R C G
Iron-sulphur centres
54NADPH? Adrenodoxin reductase
Cytochrome P?450 (mono-oxygenase)
FeS
FAD
Fe3
NADP
AH O2
A-OH H2O
NADPH H
Iron-sulphur protein
Mitochondral hydroxylase system
EMB-RCG
55E M B - R C G
EMB-RCG
56E M B - R C G
EMB-RCG
57E M B - R C G
EMB-RCG
58(No Transcript)
59(No Transcript)
60E M B - R C G
61(No Transcript)
62a-Ketoglutarate
dehydrogenase
CH2 COOH
CH2 COOH
NADHH
NAD
CH2 C COOH
CH2 C S CoA
CO2
CoA?SH
O
O
a-Ketoglutarate
Succinyl CoA
63(No Transcript)
64E M B - R C G
65Oxidative phosphorylation at the level of
respiratory chain
E M B - R C G
66E M B - R C G
67E M B - R C G
68Sequence of carriers in the respiratory chain
69Transport of reducing equivalents in
respiratory chain
E M B - R C G
70E M B - R C G
71(No Transcript)
72Reduced coenzyme Q transfers the reducing
equivalents to cytochrome b
E M B - R C G
Cytochrome b is associated with iron-sulphur
protein
The subsequent carriers,
cytochromes c1, c and a, are typical
iron-porphyrin proteins
73E M B - R C G
74(No Transcript)
75Prosthetic group of cytochrome b
Prosthetic group of cytochrome c
76E M B - R C G
77(No Transcript)
78Components of respiratory chain do not function
as discrete carriers of reducing equivalents
E M B - R C G
They are organized into four complexes
Each complex acts as a specific oxido-reductase
79E M B - R C G
80(No Transcript)
81Complex I, II, III and IV in respiratory chain
EMB-RCG
82EMB-RCG
83(No Transcript)
84(No Transcript)
85(No Transcript)
86(No Transcript)
87E M B - R C G
88(No Transcript)
89E M B - R C G
90E M B - R C G
91E M B - R C G
92E M B - R C G
93E M B - R C G
94E M B - R C G
95The ratio of ADP molecules phospho-rylated to
number of oxygen atoms reduced is known as PO
ratio
E M B - R C G
PO ratio is three when the reducing equivalents
are accepted by NAD
PO ratio is two when the reducing equivalents
are accepted by FAD
96Mechanism of oxidative phosphorylation
Mechanism by which oxidation and phospho-rylation
are coupled remained unclear for long
E M B - R C G
Several hypotheses were advanced to explain the
mechanism of this coupling
The chemiosmotic hypothesis is the most plausible
97Chemiosmotic hypothesis
Proposed by Mitchell
Inner mitochondrial membrane is impermeable to
protons (H)
E M B - R C G
Energy released during transport of electrons
is used to actively eject H from the matrix of
mitochondria
This ejection establishes an electro-chemical
gradient across the membrane
98(No Transcript)
99(No Transcript)
100E M B - R C G
101E M B - R C G
102 Re-entry of protons releases energy
103The energy released during influx of protons is
used to activate a membrane-bound enzyme
This enzyme, vectorial ATP synthetase, converts
ADP and Pi into ATP
104E M B - R C G
105EMB-RCG
106EMB-RCG
107E M B - R C G
108E M B - R C G
109 The subunits of F0 and F1 components of
vectorial ATP synthetase
EMB-RCG
110When hydrogen ions flow back into the matrix
Two hydrogen ions
combine with two electrons and one oxygen
atom of inorganic phosphate
E M B - R C G
Water is formed and inorganic phosphate is
converted into a highly reactive form
This form readily combines with ADP by a
high-energy bond
EMB-RCG
111(No Transcript)
112(No Transcript)
113(No Transcript)
114E M B - R C G
EMB-RCG
115E M B - R C G
EMB-RCG
116EMB-RCG
117(No Transcript)
118There is strong experimental evidence in support
of chemiosmotic hypothesis
Building up of H gradient is the basic premise
of the hypothesis
E M B - R C G
It is seen that on bathing mitochondria in a
fluid having a relatively high H concen-tration,
phosphorylation occurs in the absence of
oxidation
119E M B - R C G
120The PH and HO ratios of various substrates
are also in agreement with the experimental
evidence
E M B - R C G
Thus, the chemiosmotic hypothesis has now become
the accepted theory to explain oxidative
phosphorylation in the respiratory chain
121Inhibitors of oxidative phosphorylation
E M B - R C G
122E M B - R C G
123E M B - R C G
124Inhibitors of Oxidative Phosphorylation
125 Uncouplers of oxidative phosphorylation
E M B - R C G
126E M B - R C G
127An endogenous uncoupler
E M B - R C G
128(No Transcript)
129E M B - R C G
130Regulation of oxidative phosphorylation
E M B - R C G
131E M B - R C G
132Oxidation of extra-mitochondrial NADH
E M B - R C G
133Two important mechanisms are
E M B - R C G
EMB-RCG
134Glycerophosphate shuttle
E M B - R C G
135E M B - R C G
136 Glycerophosphate shuttle
137Malate shuttle
E M B - R C G
138E M B - R C G
139E M B - R C G
140E M B - R C G
141Malate shuttle
Cytosol
Mitochondrial Matrix
Oxalo-
Oxalo-
NADHH
NADHH
Aspartate
acetate
acetate
Aspartate
3
4
3
4
3
3
4
GOT
GOT
3
3
3
3
MDH
MDH
Glutamate
a
-Keto-
a-Keto-
Glutamate
3
5
glutarate
glutarate
3
4
Malate
Malate
NAD
NAD
4
142Transport across mitochondrial membrane
E M B - R C G
143(No Transcript)
144(No Transcript)
145 Transport of pyruvate and H into mitochondria
is an example of a symport
Membrane
Mitochondrial matrix
Cytosol
Pyruvate
Pyruvate
H
H
146Some important antiports
Memb
Cytosol
Matrix
ADP
ADP
ATP
ATP
Malate
Malate
a-Ketoglutarate
a-Ketoglutarate
Glutamate
Glutamate
Aspartate
Aspartate
Malate
Malate
Citrate H
Citrate H
147Thank you