Download Metabolism: Introduction

Introduction to Metabolism
Metabolism
 The sum of the chemical changes that convert
nutrients into energy and the chemically
complex products of cells
 Hundreds of enzyme reactions organized into
discrete pathways
 Substrates are transformed to products via
many specific intermediates
 Metabolic maps portray the reactions
A Common Set of Pathways
 Organisms show a marked similarity in their
major metabolic pathways
 Evidence that all life descended from a
common ancestral form
 There is also significant diversity
 Autotrophs use CO2; Heterotrophs use organic
carbon; Phototrophs use light; Chemotrophs
use Glc, inorganics use S and obtain chem
energy through food generated by
phototrophs.
The Sun is Energy for Life
Phototrophs use light to drive synthesis
of organic molecules
Heterotrophs use these as building
blocks
CO2, O2, and H2O are recycled
Metabolism
 Metabolism consists of catabolism and
anabolism
 Catabolism: degradative pathways
 Usually energy-yielding!
 “destructive metabolism”
 FUELS -> -> CO2 + H2O + useful energy
 Anabolism: biosynthetic pathways
 energy-requiring!
 “constructive metabolism”
 Useful energy + small molecules --> complex molecules
Organization in Pathways
 Pathways consist of sequential steps
 The enzymes may be:
 Separate
 Form a multienzyme complex
 A membrane-bound system
 New research indicates that multienzyme
complexes are more common than once
thought
Catabolism and Anabolism
 Catabolic pathways converge to a few end
products
 Anabolic pathways diverge to synthesize many
biomolecules
 Some pathways serve both in catabolism and
anabolism and are called amphibolic pathways
Digestion of food polymers:
 enzyme-catalyzed hydrolysis
Glycolysis:
 glucose catabolism
 generate ATP without consuming oxygen (anaerobic)
Citric Acid Cycle:
 metabolism of acetyl-CoA derived from pyruvate, fatty
acids, and amino acids
 acetyl oxidized to CO2
 operates under aerobic conditions
 reduction of coenzymes NAD+ and FAD; energy used to
produce ATP
Oxidative phosphorylation:
 reduction of molecular oxygen by NADH and FADH2
 energy of reduced compounds used to pump protons
across a cell membrane
 potential energy of electrochemical gradient drives
phosphorylation of ADP to ATP
Comparing Pathways
Anabolic & catabolic pathways involving
the same product are not the same
Some steps may be common to both
Others must be different - to ensure that
each pathway is spontaneous
This also allows regulation mechanisms
to turn one pathway and the other off
METABOLIC REGULATION
Regulated by controlling:
1. Amounts of enzymes
2. Catalytic activities
3. Accessibility of substrates
The ATP Cycle
 ATP is the energy currency of cells
 In phototrophs, light energy is transformed
into the light energy of ATP
 In heterotrophs, catabolism produces ATP,
which drives activities of cells
 ATP cycle carries energy from photosynthesis
or catabolism to the energy-requiring
processes of cells
Redox in Metabolism
 NAD+ collects electrons released in catabolism
 Catabolism is oxidative - substrates lose
electrons, usually H- ions
 Anabolism is reductive - NADPH provides the
electrons for anabolic processes, and the
substrates gain electrons
WHY ATP?
Free energy is released when ATP is
hydrolyzed.
This energy drives reactions that need it
(eg. muscle contraction)
Recall coupled reactions
ATP has a higher phosphoryl transfer
potential
RECURRING MOTIFS IN METAB
Certain compounds keep on recurring or
appearing in metabolic reactions and
their functions are the same in the
processes
Metab looks complicated but reactions
are actually limited and repeating.
ACTIVATED CARRIERS
These species help carry out the
metabolic reactions, even nonfavorable
ones, at times
Example: ATP (activated carrier of
phosphoryl groups)
Activated carriers of electrons for
fuel oxidation: e- acceptors!
 Aerobic systems: O2 is the final eacceptor, but this does not occur
directly
 Fuels first transfer e- to carriers:
pyridine molecules or flavins.
NAD+:
nicotinamide
adenine
dinucleotide
Activated carriers of electrons for
fuel oxidation: e- acceptors!
FAD: Flavin
adenine
dinucleotide
Activated carrier of electrons for
reductive biosynthesis: e- donors!
NADPH: common
electron donor
R is phosphate
group
Activated carrier of two-carbon
fragments
COENZYME A: carrier of acyl groups
Activated carrier of two-carbon
fragments
VITAMINS
Many vitamins are "coenzymes" molecules that bring unusual chemistry
to the enzyme active site
Vitamins and coenzymes are classified
as "water-soluble" and "fat-soluble"
The water-soluble coenzymes exhibit the
most interesting chemistry
Key Reactions in Metabolism
1. REDOX reactions
Electron carriers are needed!
2. LIGATION reactions
 Bond formation facilitated by ATP cleavage
3. ISOMERIZATION reactions
4.GROUP TRANSFER
5.HYDROLYTIC reactions
Bond cleavage by addition of H2O
6.ADDITION of functional groups
to double bonds or REMOVAL of
groups to form double bonds
Uses lyases
GLYCOLYSIS
Glycolysis
1897: Hans and Eduard Buchner
(Sucrose cell-free experiments;
fermentation can take place outside of
living cells) METABOLISM became simple
chemistry
Glycolysis: “Embden-Meyerhof pathway”
The all-important Glucose
The only fuel the brain uses in nonstarvation conditions
The only fuel red blood cells can use
WHY?
 Evolutionary: probably available for
primitive systems
The products and their fates
Glycolysis
AKA Embden-Meyerhof-Parnas Pathway
Involves the oxidation of glucose
Products:
 2 Pyruvate
 2 ATP
 2 NADH
Cytosolic
Glycolysis
Anaerobic
The entire
process does not
require O2
Glycolysis: General Functions
Provide energy in the form of ATP
Generate intermediates for other
pathways:
 Hexose monophosphate pathway
 Glycogen synthesis
 Pyruvate dehydrogenase
 Fatty acid synthesis
 Krebs’ Cycle
 Glycerol-phosphate (TG synthesis)
Specific functions of glycolysis
 Red blood cells (RBCs)
 Rely exclusively for energy
 Skeletal muscle
 Source of energy during exercise, particularly high
intensity exercise
 Adipose tissue
 Source of glycerol-P for TG synthesis
 Source of acetyl-CoA for FA synthesis
 Liver
 Source of acetyl-CoA for FA synthesis
 Source of glycerol-P for TG synthesis
Regulation of Cellular Glucose Uptake
 Brain & RBC:
 The GLUT-1 transporter has high affinity for
glucose and is always saturated.
 Ensures that brain and RBC always have glucose.
 Liver:
 The GLUT-2 glucose transporter has low affinity
and high capacity.
 Uses glucose when fed at rate proportional to glucose
concentration
 Muscle & Adipose:
 The GLUT-4 transporter is sensitive to insulin
Glucose Utilization
Phosphorylation of glucose
 Commits glucose for use by that cell
 Energy consuming
Hexokinase: muscle and other tissues
Glucokinase: liver
Properties of Glucokinase and
Hexokinase
Regulation of Cellular Glucose
Utilization in the Liver
 Feeding




Blood glucose concentration high
GLUT-2 taking up glucose
Glucokinase induced by insulin
High cell glucose allows GK to phosphorylate
glucose for use by liver
 Post-absorptive state




Blood & cell glucose low
GLUT-2 not taking up glucose
Glucokinase not phophorylating glucose
Liver not utilizing glucose during post-absorptive
state
Regulation of Cellular Glucose
Utilization in the Liver
Starvation
 Blood & cell glucose concentration low
 GLUT-2 not taking up glucose
 GK synthesis repressed
 Glucose not used by liver during starvation
Regulation of Cellular Glucose
Utilization in the Muscle
 Feeding and at rest




High blood glucose, high insulin
GLUT-4 taking up glucose
HK phosphorylating glucose
If glycogen stores are filled, high G6P inhibits HK,
decreasing glucose utilization
 Starving and at rest




Low blood glucose, low insulin
GLUT-4 activity low
HK constitutive
If glycogen stores are filled, high G6P inhibits HK,
decreasing glucose utilization
Regulation of Cellular Glucose
Utilization in the Muscle
Exercising Muscle (fed or starved)
 Low G6P (being used in glycolysis)
 No inhibition of HK
 High glycolysis from glycogen or blood
glucose
Regulation of Glycolysis
Regulation of 3 irreversible steps
PFK-1 is rate limiting enzyme and
primary site of regulation.
Regulation of
Glycolysis
Most important
regulation hub!
Regulation of PFK-1 in Muscle
 Relatively constitutive
 Allosterically stimulated by AMP
 High glycolysis during exercise
 Allosterically inhibited by
 ATP
 High energy, resting or low exercise
 Citrate
 Build up from Krebs’ cycle
 May be from high FA beta-oxidation -> hi acetyl-CoA
 Energy needs low and met by fat oxidation
Regulation of PFK-1 in Liver
Inducible enzyme
 Induced in feeding by insulin
 Repressed in starvation by glucagon
Allosteric regulation
 Like muscle w/ AMP, ATP, Citrate
 Activated by Fructose-2,6-bisphosphate
Fermentation
Anaerobic respiration!
Produces ATP without oxygen.
No ETC is present since there is no
oxygen
NAD+ gets recycled by use of an organic
hydrogen acceptor like lactate or
ethanol.
Common in prokaryotes and very useful
to humans.
Fermentation
Two type lactic acid and alcohol
fermentation.
A build up of lactate in your muscles
from over exerting yourself and not
taking in enough oxygen causes
soreness.
Alcohol fermentation has a by product of
CO2 and ethanol which is used to make
alcoholic beverages. Yeast and fungus
go through alcohol fermentation.
The release of CO2 by yeast is what
causes bread to rise.
Alcohol Fermentation
 pyruvate is
converted to
ethanol in two
steps.
 Alcohol
fermentation
by yeast is
used in
brewing and
winemaking.
Lactic Acid Fermentation
 pyruvate is reduced
directly by NADH to form
lactate
 Lactic acid fermentation by
some fungi and bacteria is
used to make cheese and
yogurt
 The waste product,
lactate, may cause
muscle fatigue, but
ultimately it is
converted back to
pyruvate in the liver.
The Tricarboxylic Acid (TCA) Cycle
 Also known as the Krebs Cycle and Citric Acid
Cycle
 The citric acid cycle is the final common
pathway for the oxidationof fuel molecules:
amino acids, fatty acids, & carbohydrates.
 Most fuel molecules enter the cycle as acetyl
coenzyme A
 This cycle is the central metabolic hub of the
cell
The Tricarboxylic Acid (TCA) Cycle
 The citric acid cycle oxidizes two-carbon units
 Entry to the cycle and metabolism through it
are controlled
 It is the gateway to aerobic metabolism for
any molecule that can be transformed into an
acetyl group or dicarboxylic acid,
 It is also an important source of precursors for
building blocks
Overview of the TCA Cycle
1. The function of the cycle is the harvesting of highenergy electrons from carbon fuels
2. The cycle itself neither generates ATP nor includes O2
as a reactant
3. Instead, it removes electrons from acetyl CoA & uses
them to form NADH & FADH2 (high-energy electron
carriers)
4. In oxidative phosphorylation, electrons from
reoxidation of NADH & FADH2 flow through a series of
membrane proteins (electron transport chain) to
generate a proton gradient
Overview of the TCA Cycle
5. These protons then flow back through ATP
synthase to generate ATP from ADP &
inorganic phosphate
6. O2 is the final electron acceptor at the end of
the electron transport chain
7. The cytric acid cycle + oxidative
phosphorylation provide > 95% of energy
used in human aerobic cells
Fuel for the Citric Acid Cycle
Pantothenate
Thioester bond
to acetate
-mercapto-ethylamine
Mitochondrion
70
Double membrane, & cristae: invaginations of inner membrane
Mitochondrion
Oxidative
decarboxilation
of pyruvate, & citric acid
cycle take place in the
matrix, along with fatty
acid oxidation
Site of oxidative
phosphorylation
Permeable
TCA Cycle: Overview
Input: 2-carbon units
in the form of AcetylCoA
Output: 2 CO2, 1 GTP,
& 8 high-energy
Electrons in the form
of reducing elements
Cellular Respiration
8 high-energy
electrons from
carbon fuels
Electrons reduce
O2 to generate a
proton gradient
ATP synthesized
from proton
gradient
Acetyl-CoA: Link between glycolysis and TCA
Acetyl CoA is the
fuel for the citric acid
cycle
Pyruvate Dehydrogenase:
 AKA PDH
 The enzyme that links glycolysis with other pathways
 Pyruvate + CoA + NAD -> AcetylCoA + CO2 + NADH
The PDH Complex
Multi-enzyme complex
 Three enzymes
 5 co-enzymes
 Allows for efficient direct transfer of product from
one enzyme to the next
The PDH Reaction
 E1: pyruvate dehydrogenase
 Oxidative decarboxylation of pyruvate
 E2: dihydrolipoyl transacetylase
 Transfers acetyl group from TPP to lipoic acid
 E3: dihydrolipoyl dehydrogenase
 Transfers acetly group to CoA, transfers electrons from reduced
lipoic acid to produce NADH
Regulation of PDH
Muscle
 Resting (don’t need)
 Hi energy state
 Hi NADH & AcCoA
 Inactivates PDH
 Hi ATP & NADH & AcCoA
 Inhibits PDH
 Exercising (need)
 Low NADH, ATP, AcCoA
Regulation of PDH
Liver
Fed (need to make
FA)
 Hi energy
 Insulin activates PDH
Starved (don’t need)
 Hi energy
 No insulin
 PDH inactive
Coenzymes
Vitamin B1
FAD
FAD
FADH2
NAD
Step 1: Citrate formation
Enzyme: Citrate synthase
Condensation reaction
Hydrolysis reaction
Step 2: Isomerization of citrate to isocitrate
Enzyme: Aconitase
Dehydration
Hydration
Step 3: Isocitrate to α-ketoglutarate
Enzyme: Isocitrate dehydrogenase
1st NADH produced!
1st CO2 removed
Step 4: Succinyl-CoA formation
Enzyme: α-ketoglutarate dehydrogenase
2nd NADH produced!
2nd CO2 removed!
Step 5: Succinate formation
Enzyme: Succinyl CoA synthetase
GTP produced
• Equivalent to ATP!
• GTP + ADP  GDP + ATP
Step 6: Succinate to Fumarate
Enzyme: Succinate dehydrogenase
FADH2 produced!
Step 7: Fumarate to Malate
Enzyme: Fumarase
Step 8: Malate to Oxaloacetate
Enzyme: Malate dehydrogenase
3rd NADH produced
The TCA Cycle
Summary of the Reactions in TCA
Control of the TCA Cycle
 Regulated primarily by
ATP & NADH concentrations
 control points:
 Pyruvate
dehydrogenase
 isocitrate
dehydrogenase
 - ketoglutarate
dehydrogenase
Biosynthetic roles of the TCA cycle
OXIDATIVE PHOSPHORYLATION
What’s the
point?
The point
is to make
ATP!
ATP
2006-2007
ATP accounting so far…
Glycolysis  2 ATP
Kreb’s cycle  2 ATP
Life takes a lot of energy to run, need to
extract more energy than 4 ATP!
What’s the
point?
A working muscle recycles over
10 million ATPs per second
There is a better way!
Electron Transport Chain
 series of molecules built into inner
mitochondrial membrane
 along cristae
 transport proteins & enzymes
 transport of electrons down ETC linked to
pumping of H+ to create H+ gradient
 yields ~30-32 ATP from 1 glucose!
 only in presence of O2 (aerobic respiration)
That
sounds more
like it!
O2
Mitochondria
Double membrane
 outer membrane
 inner membrane
 highly folded cristae
 enzymes & transport
proteins
 intermembrane space
 fluid-filled space between
membranes
Oooooh!
Form fits
function!
Electron Transport Chain
Inner
mitochondrial
membrane
Intermembrane space
C
Q
NADH
dehydrogenase
cytochrome
bc complex
Mitochondrial matrix
cytochrome c
oxidase complex
Remember the Electron Carriers?
Glycolysis
glucose
Krebs cycle
G3P
4 NADH
Time to
break open
the bank!
8 NADH
2 FADH2
Electron Transport Chain
Inner
mitochondrial
membrane
Intermembrane space
C
Q
NADH
dehydrogenase
cytochrome
bc complex
Mitochondrial matrix
cytochrome c
oxidase complex
But what “pulls” the
electrons down the ETC?
O2
electrons
flow downhill
to O2
oxidative phosphorylation!
Electrons flow downhill
Electrons move in steps from
carrier to carrier downhill to O2
 each carrier more electronegative
 controlled oxidation
 controlled release of energy
make ATP
instead of
fire!
We did it!
“proton-motive” force
H+
H+
Set up a H+
gradient
H+
H+
H+
H+
H+
H+
Allow the protons
to flow through
ATP synthase
Synthesizes ATP
ADP + Pi  ATP
Are we
there yet?
ADP + Pi
ATP
H+
Chemiosmosis
 The diffusion of ions across a membrane
 build up of proton gradient just so H+ could flow
through ATP synthase enzyme to build ATP
Chemiosmosis
links the
Electron
Transport Chain
to ATP synthesis
So that’s
the point!
Peter Mitchell
Proposed chemiosmotic hypothesis
 revolutionary idea at the time
True story.
proton motive force
1920-1992
Pyruvate from
cytoplasm
Inner
+
mitochondrial H
membrane
H+
Intermembrane
space
Electron
transport
C system
Q
NADH
Acetyl-CoA
2. Electrons
provide energy
1. Electrons are harvested to pump protons
and carried to the transport
across the
system.
membrane.
-
NADH
Krebs
cycle
e-
e
FADH2
e-
ATP
Mitochondrial
matrix
e-
H2O
3. Oxygen joins
with protons to
form water.
1 O
2 +2
2H+
O2
H+
CO2
2
H+
32 ATP
4. Protons diffuse back in
down their concentration
gradient, driving the
synthesis of ATP.
H+
ATP
synthase
Cellular respiration
2 ATP
+ ~2 ATP + 2 ATP + ~34 ATP
Cellular respiration
Pathway
Glycolysis
Substrate-Level
Oxidative
Phosphorylation Phosphorylation
2 ATP
2 NADH = 4 - 6
ATP
Total
ATP
6-8
CoA
2 NADH = 6 ATP
6
Krebs Cycle
2 ATP
6 NADH = 18
ATP
2 FADH2 = 4 ATP
24
TOTAL
4 ATP
32 ATP
36 - 38
Summary of cellular respiration
Oxidative phosphorylation is the process
of making ATP from the reducing
elements NADH and FADH2, with the
help of O2 and the electron transport
chain
The electron transport chain is the
structural complex that enables
oxidative phosphorylation to take place
Summary of cellular respiration
C6H12O6 + 6O2
 6CO2 + 6H2O + ~40 ATP
 Where did the glucose come from?
 Where did the O2 come from?
 Where did the CO2 come from?
 Where did the CO2 go?
 Where did the H2O come from?
 Where did the ATP come from?
 What else is produced that is not listed
in this equation?
 Why do we breathe?
Taking it beyond…
What is the final electron acceptor in
Electron Transport Chain?
O2
 So what happens if O2 unavailable?
 ETC backs up
nothing to pull electrons down chain
 NADH & FADH2 can’t unload H

 ATP production ceases
 cells run out of energy
 and you die!
WHOA!