BIOC 460 - DR.

TISCHLER LECTURE 25

OXIDATIVE PHOSPHORYLATION:
AN EXPLOSION OF ENERGY

1.

What effect does insulin have on glucose transport increases by moving transporters to the membrane

Quiz 8

2. Name one tissue where insulin controls glucose transport muscle OR adipose. 3. Glucose increases insulin secretion by causing an increase in the cell concentration of what molecule? Calcium 4. What vitamin is the precursor for NADH? Niacin 5. How is pyruvate kinase structurally modified after glucagon binds to the liver cell? Phosphorylation 6. In the red cell, which lacks mitochondria, how is NAD for glycolysis replenished to keep glycolysis going? Conversion to lactate or LDH 7. Identify an allosteric inhibitor of the phosphofructokinase -1 reaction. ATP or citrate or H+ 8. Identify an allosteric activator of the phosphofructokinase -1 reaction. AMP or fructose-2,6-bisphosphate
2

OBJECTIVES
1. Distinguish between electroneutral and electrogenic transport; describe the significance of electrogenic transport for the adenine nucleotide transporter and the role of this transporter. 2. Discuss oxidative phosphorylation in relation to the chemiosmotic model. 3. Identify the components of the ATP synthase complex, and describe their roles. 4. Explain how the malate-aspartate and the -glycerol phosphate electron shuttles generate energy from the NADH produced by glycolysis. (Do not memorize the layout of the shuttle to reproduce it but understand their key aspects) 5. Define respiratory control and uncoupling, and describe the physiological importance of these processes.

PHYSIOLOGICAL PREMISE Why do some snake and spider venoms cause cell death at the site of the bite in particular? Some of these venoms contain enzymes called phospholipases. Phospholipases hydrolyze membrane phospholipids to release fatty acids. Phospholipases in snake or spider venom degrade phospholipids in the mitochondrial membrane. The fatty acids released act as natural uncouplers that, as is described in this lecture, prevent oxidative phosphorylation by destroying the pH gradient. Consequently the cell dies because of an inability to produce enough energy.

MITOCHONDRIAL TRANSPORT SYSTEMS Examples of Electroneutral Transport: Pyruvate1- moves into matrix and OH1- moves out Phosphate1- moves into matrix and OH1- moves out

Citrate3- + H+ exchanges with malate2-

MITOCHONDRIAL TRANSPORT SYSTEMS Inner Intermembrane Membrane Space Matrix Side ATP4ADP3Net negative charge moves out

Adenine nucleotide translocase

Electrogenic Transport

Glu1- + H+

Asp1-

Aspartate translocase

Figure 1. Electrogenic transport system in mitochondria.

33- ATP4OH- ADP ADP

INTERMEMBRANE SPACE 4H+ complex I NADH FMNH2 + H+4H+ eeeinner e membrane NAD+ CoQ complex III + 4H 2H e cyt b e2H+ C1 O2 e cyt e-H O e a-a3 -C 2 e 4H+ complex IV

OH-

ATP43ADP ADP3-

3OH- ADP ADP 3-

ATP4-

Pi

3ADP3-

3H+

ATP4-

F1

MATRIX

stalk

Fo
Proton gradient/ Charge gradient 3H+

Figure 2. Oxidative Phosphorylation

FEATURES OF THE CHEMIOSMOTIC MODEL:

mitochondrial inner membrane is impermeable to protons by simple diffusion measurable proton (pH) gradient exists across the inner membrane collapse of the proton gradient (uncoupling) abolishes ATP synthesis but accelerates O2 consumption inhibition of the respiratory chain prevents ATP synthesis because pumping of protons ceases

e = electrons Glucose NAD+

INNER OUTER MEMBRANE GLYCOLYSIS eMEMBRANE Pyruvate

Complex I

eOAA Glu0 NADH e-

NADH e(1) NAD+

OAA (6)

Glu0 Asp-1 KG

(5)

Asp-1

(4)
(3) eMalate NAD+ MATRIX

KG (2)

eMalate CYTOPLASM

Figure 3. The malate-aspartate shuttle.

e = electrons CYTOPLASM Glucose GLYCOLYSIS NAD+ Pyruvate Dihydroxyacetone phosphate (DHAP) (1) Glycerol 3-phosphate OUTER MEMBRANE

O2
e CoQ e FADH2

INNER MEMBRANE

MATRIX

NADH e NAD+

DHAP (2) G3P

e Glycerol-3-phosphate dehydrogenase

FAD

Figure 4. Glycerol phosphate shuttle. Cytoplasmic glycerol 3phosphate dehydrogenase (1) oxidizes NADH. Glycerol 3-phosphate dehydrogenase in the inner membrane (2) reduces FAD to FADH2.

RESPIRATORY CONTROL depends on the availability of ADP increased ADP in matrix opens proton channel

protons move through channel down pH gradient

respiration increases to compensate for decline in pH gradient; oxygen consumption (respiration) controlled at low ADP, ATP synthesis ceases, pH gradient builds up, oxygen use diminishes as ATP needs rise (i.e., ADP increases) respiration is again accelerated inhibition of respiratory chain causes loss of control

UNCOUPLER EFFECTS hydrophobic molecules that bind protons take protons into matrix to collapse pH gradient without pH gradient, synthesis of ATP ceases electron transport chain operates at high rate; protons are pumped out rapidly in attempt to restore pH gradient energy is released as heat and the body temperature rises respiratory control is lost uncoupled brown fat mitochondria generates body heat in infants until shivering reflex develops

LACTATE

GLYCOGEN
NADH

GLUCOSE
NADH

CYTOPLASM
ALANINE

+ nitrogen INTERMEMBRANE SPACE

PYRUVATE MALATE-ASP SHUTTLE

complex IV

Mal

PYRUVATE PDH Oaa ACETYL CoA MDH CS

PC

H+

TCA CYCLE SDH

NADH

ICDH
KgDH

MITOCHONDRIAL MATRIX

complex III

NAD ADP + Pi
complex I

H+

ATP

X
uncoupler
H+ ATP

Fig. 5 Effects of uncoupling on glucose, lactate and mitochondrial metabolism

ADP

Pi

Medical Scenario I: As a current medical student and a former lawyer you become interested in what could have been a lucrative malpractice case in the 1920's. At that time, some physicians, without the benefit of modern biochemistry knowledge, prescribed dinitrophenol as a weight-reducing agent. For some patients it worked satisfactorily, the only side effects being sweating and a slight elevation in temperature. However, in patients who were more overweight and required long-term treatments, their body temperature increased to such an extent that death resulted. Associated with their condition was a marked increase in their rate of respiration and an inability to gain weight even with increased caloric intake. Medical Scenario II:

During World War II workers in munitions factories were exposed to a chemical called trinitrophenol. A common occurrence in these workers was frequent absenteeism due to elevated temperature, weakness and accelerated breathing that improved after several days at home. What would you have recommended to reduce the incidence of such illness?