Post on 30-Mar-2020
Lecture Notes for
Chapter 15 Oxidative Phosphorylation
Essential Biochemistry Third Edition
Charlotte W. Pratt | Kathleen Cornely
Copyright © 2014 John Wiley & Sons, Inc. All rights reserved.
15-1 The Thermodynamics of Oxidation-Reduction Reactions
oxidant reductant
還原 = 獲得電子 = 與氫鍵結增加
Reduction potential indicates a substance’s
tendency to accept electrons
Half reaction
將還原電位量化 還原 = 分子接受電子 電位 = 伏特的改變 還原電位 = 分子接受電子後伏特的改變
The free energy change can be
calculated from the change in
reduction potential
Rule
• 不同分子有不同還原電位
• 還原電位高 = 分子拉電子力強
• Δ ε °’ = ε °’ acceptor – ε °’
donor
• 為方便記憶,均以獲得電子方向
之還原電位進行計算
• Donor : NADH,ε °’ = -0.315
• Acceptor : Q,ε °’ = +0.045
• 故Δ ε °’ =0.045 – (-0.315) =
0.36
• 淨反應如下:
The actual reduction potential depends on
the actual concentrations of oxidized and
reduced species.
The Nernst Equation
R = Gas Constant = 8.3145 J‧mol-1‧K-1 T = temperature in Kelvin
n = # of electrons F = Faraday’s constant = 96,485 J‧V-1‧K-1
實際還原電位受氧化態與還原態濃度影響,以Nernst equation計算之,其中:
換算為自由能:
亦即還原電位為正值時,Δ G°’為負值,可自發之放熱反應
Δ ε °’= +0.360
Δ ε °’= +0.190
Δ ε °’= +0.580
15-2 Mitochondrial Electron Transport
• Respiration: 還原能最終傳遞至氧氣,使其還原為H2O,因此消耗氧氣而獲得能量,稱之為呼吸作用
Electron transport takes place in the
mitochondrion.
© 2014 John Wiley & Sons, Inc. All rights reserved.
粒線體結構 可通透
不可通透
生產ATP所必需原料: • NADH • ADP • Pi
The malate-aspartate shuttle
transports reducing agents across
the inner mitochondrial membrane.
何種運送機制可使NADH由粒線體外轉移至粒線體內?
NADH除了由TCA生產,亦可由粒線體外轉換而補充:
A different transport system is used to
move ATP from the matrix to the cytosol.
ATP translocase
:負責ATP換出,ADP換入
Symport protein
:利用H+的濃度差同向運送Pi進入
Oxidative phosphorylation
© 2014 John Wiley & Sons, Inc. All rights reserved.
Complex I 接受來自NADH之電子,排出H+
Complex II 接受來自FADH2之電子
Complex III 匯整電子流,排出H+
Complex IV 將O2還原為水,排出H+
Complex I binds ubiquinone.
• Complex I 結構解析
– 使電子由NADH轉移到輔酶Q
– 主要利用FMN(B2)與Fe-S 為輔因子
– 嵌在粒線體內膜中,故結構上有許多α -helix二級結構
© 2014 John Wiley & Sons, Inc. All rights reserved.
FMN can pick up two electrons from
NADH.
© 2014 John Wiley & Sons, Inc. All rights reserved.
第一步:FMN由NADH+H+獲得2電子形成FMNH2
Iron-sulfur clusters
undergo
one-electron
transfer reactions.
© 2014 John Wiley & Sons, Inc. All rights reserved.
第二步:一系列的Fe-S輔因子協助電子傳送
第三步:輔酶Q接過2電子,形成QH2
Proton wire (質子線路)
a series of hydrogen-bonded protein groups plus water molecules that form a
chain through which a proton can be rapidly relayed.
Complex I Function
移出2H+
移出2H+
共計4H+
Structure of Complex II
(succinate dehydrogenase)
Succinate dehydrogenase,即為TCA循環中的一個產生FADH2之enzyme FAD接收 Succinate之電子,產生FADH2
FADH2之電子經由Fe-S傳至CoQ使其還原為CoQH2
但FADH2還原電位幾乎為0,故淨反應放熱不足以產生ATP,亦無法排出質子
Complex I電子來自____,而Complex II電子來自____,主要都傳給______
TCA循環中哪一個酶與氧化磷酸化有關?
Complex III transfers electrons from
ubiquinol to cytochrome c
Heme prosthetic group One-electron reduction
Cytochrome 細胞色素
• 利用含鐵血紅素(Heme b)輔基傳送電子之蛋白
• 鐵離子可於還原態Fe2+與氧化態Fe3+切換,傳送一個電子
• QH2有兩個電子,故需要 2 Cytochrome c
cytochrome b
Iron–Sulfur Protein
(2Fe–2S cluster)
cytochrome c1
Structure of Mammalian Complex III
The Q cycle
• 由QH2傳進2個電子,但cytochrome c一次只能收1個電子,故由cytochrome b保留,再傳回Q形成‧Q-
• 另一個QH2再傳進2個電子
,由 cytochrome b再傳至‧Q-,與2H+形成新的QH2
移出2H+ (屬於I)
移出2H+
移出2H+ 共計4H+
Summary for Complex III function
• 每Round釋出2H+,兩Round釋出4H+,運送2電子
Complex IV oxidizes cytochrome c
and reduces O2.
• Cytochrome c 一次轉移1個電子至Complex IV
• Complex IV 將O2還原為水,使Cytochrome c氧化,故又稱為Cytochrome c oxidase
© 2014 John Wiley & Sons, Inc. All rights reserved.
氧化磷酸化反應中,最終的電子接受者為何?
More on Complex IV Function
• 1NADH轉移2個電子,最終使Cyto c轉移2個質子出去
• 另外2個質子與氧結合,
½ O2 H2O
© 2014 John Wiley & Sons, Inc. All rights reserved.
質子梯度(Proton gradient)的形成
© 2014 John Wiley & Sons, Inc. All rights reserved.
Complex I排出4質子 Complex III排出4質子
1分子NADH+H+ 提供2電子
Complex IV排出2質子
[H+]↑,pH↓
[H+]↓,pH↑
1分子NADH參與呼吸鏈,可由粒線體基質排出多少質子?
15-3 Chemiosmosis
ADP + Pi ATP
Δ G°’= +30.5kJ/mol
如何轉移能量形成ATP?
Chemiosmosis links electron transport
and oxidative phosphorylation
• Chemiosmotic theory 化學滲透假說:
粒線體內膜內外的質子梯度 proton gradient 推動反應
• Proton-motive force :
質子梯度差趨使質子由高濃度往低濃度移動,產生之動能進而合成ATP
Computing the free energy change
for the imbalance of protons.
化學能推動 電位差推動
Z = Ion’s Charge
Δy = Membrane Potential
© 2014 John Wiley & Sons, Inc. All rights reserved.
整合
15-4 ATP Synthase (Complex V)
ATP synthase
F1球體:伸進粒線體基質
F0 跨膜結構
ATP synthase rotates as it translocates
protons
a subunit
b subunit
c subunit X 12
F1:α *3, β *3, γ *1, δ *1, ε *1
ATP synthase rotates as it
translocates protons.
• H+ 由a subunit通道進入c subunit.
• 結構誘導H+單方向轉動,帶動整個c subunit單方向旋轉
• 每個c subunit有1個H+
結合位,1個H+進入即帶動旋轉1個c subunit而釋出1個H+
© 2014 John Wiley & Sons, Inc. All rights reserved.
ATP synthase rotates as it translocates
protons
1個質子轉動30°,4個質子轉動1/3圈,帶動γ subunit轉動120°
γ rotates in steps of 120° 4 proton = 1 ATP
僅β subunit具催化活性
The binding change mechanism explains
how ATP is made
• Binding change mechanism: 隨著γ subunit的轉動,αβ subunit有三種構型,要ADP與Pi結合才使結構得以旋轉,稱之Binding change mechanism
• Loose-binding (L): 疏鬆結構,此時ADP與Pi可就定位
• Tight-binding (T): 緊密結構,此時將ADP與Pi壓縮為ATP
• Open (O): 開放結構,此時ATP釋出
ADP與Pi於α β subunit處於何種狀態才可接上ATP synthase?
O: Open T: Tight L: Loose
Step 1 Loose state ADP and Pi bind to β
Step 2 Tight state ADP+Pi → ATP
Step 3 Open state ATP release
© 2014 John Wiley & Sons, Inc. All rights reserved.
Quantifying Oxidative Phosphorylation
• 氧化磷酸化反應,以P:O敘述氧消耗與磷酸化的比例
• P:O ratio = # phosphorylation of ADP per # of oxygen
atoms reduced (每個氧原子還原使ADP磷酸化比例)
– 1原子氧還原等於產生10個H+
– 1ADP磷酸化需要4個H+
– 同樣以H+計算,則使氧消耗與磷酸化得到共同的換算依據
© 2014 John Wiley & Sons, Inc. All rights reserved.
• ADP phosphorylation ∝ Oxygen atoms reduced
1 NADH (2e-)
→Complex I(4H+ out)
→Complex III(4H+ out)
→Complex IV(2H+ out) and ½ O2 reduced
4H+ = 1ATP, 10H+ = 2.5 ATP, Oxygen atom = 1
• So the P:O ratio of 1NADH = 2.5
The P:O ratio describes the stoichiometry of
oxidative phosphorylation
The P:O ratio describes the stoichiometry of
oxidative phosphorylation
• 1 QH2 (2e-)
→Complex II
→Complex III(4H+ out)
→Complex IV(2H+ out) and ½ O2 reduced
4H+ = 1ATP, 6H+ = 1.5 ATP, Oxygen atom = 1
So the P:O ratio of 1QH2 = 1.5
一分子NADH/QH2分別可產生多少ATP? NADH/NADPH/QH2/FADH2何者不使用於呼吸鏈反應?
The rate of oxidative phosphorylation
depends on the rate of fuel catabolism
•Reduced cofactors - (NADH and QH2) 不足
•Binding change mechanism – ADP與Pi不足
•pH調控之ATP synthase inhibitor
•基質內pH值偏高→失去與ATP synthase結合之能力→質子濃度差促進ATP合成