Photosynthesis

Chapter 8
 Photosynthesis Overview
Energy for all life on Earth ultimately comes
from photosynthesis.

6CO2 + 12H2O C6H12O6 + 6H2O + 6O2

Oxygenic photosynthesis is carried out by:

cyanobacteria, 7 groups of algae,
all land plants

2
 Photosynthesis Overview
Photosynthesis is divided into:
light-dependent reactions
-capture energy from sunlight
-make ATP and reduce NADP+ to NADPH
carbon fixation reactions
-use ATP and NADPH to synthesize
organic molecules from CO2

3
4
 Photosynthesis Overview
Photosynthesis takes place in chloroplasts.

thylakoid membrane – internal membrane

arranged in flattened sacs
-contain chlorophyll and other pigments

grana – stacks of thylakoid membranes

stroma – semiliquid substance surrounding
thylakoid membranes
5
6
 Discovery of Photosynthesis
The work of many scientists led to the
discovery of how photosynthesis works.

Jan Baptista van Helmont (1580-1644)

Joseph Priestly (1733-1804)
Jan Ingen-Housz (1730-1799)
F. F. Blackman (1866-1947)

7
 Discovery of Photosynthesis
C. B. van Niel, 1930’s
-proposed a general formula:
CO2+H2A + light energy CH2O + H2O + 2A
where H2A is the electron donor
-van Niel identified water as the source of the
O2 released from photosynthesis
-Robin Hill confirmed van Niel’s proposal that
energy from the light reactions fuels carbon
fixation 8
 Pigments
photon: a particle of light
-acts as a discrete bundle of energy
-energy content of a photon is inversely
proportional to the wavelength of the light
photoelectric effect: removal of an electron
from a molecule by light
-occurs when photons transfer energy to
electrons
9
10
 Pigments
Pigments: molecules that absorb visible
light

Each pigment has a characteristic

absorption spectrum, the range and
efficiency of photons it is capable of
absorbing.

11
12
 Pigments
chlorophyll a – primary pigment in plants
and cyanobacteria
-absorbs violet-blue and red light

chlorophyll b – secondary pigment

absorbing light wavelengths that
chlorophyll a does not absorb

13
14
15
 Pigments
accessory pigments: secondary pigments
absorbing light wavelengths other than
those absorbed by chlorophyll a
-increase the range of light wavelengths that
can be used in photosynthesis
-include: chlorophyll b, carotenoids,
phycobiloproteins
-carotenoids also act as antioxidants
16
 Photosystem Organization
A photosystem consists of
1. an antenna complex of hundreds of
accessory pigment molecules
2. a reaction center of one or more
chlorophyll a molecules

Energy of electrons is transferred through

the antenna complex to the reaction
center.
17
18
19
 Light-Dependent Reactions
In sulfur bacteria, only one photosystem is
used for cyclic photophosphorylation
1. an electron joins a proton to produce
hydrogen
2. an electron is recycled to chlorophyll
-this process drives the chemiosmotic
synthesis of ATP

20
21
 Light-Dependent Reactions
In chloroplasts, two linked photosystems are
used in noncyclic photophosphorylation
1. photosystem I
-reaction center pigment (P700) with a peak
absorption at 700nm
2. photosystem II
-reaction center pigment (P680) has a peak
absorption at 680nm
22
 Light-Dependent Reactions
Photosystem II acts first:
-accessory pigments shuttle energy to the
P680 reaction center
-excited electrons from P680 are transferred
to b6-f complex
-electron lost from P680 is replaced by an
electron released from the splitting of
water
23
 Light-Dependent Reactions
The b6-f complex is a series of electron
carriers.
-electron carrier molecules are embedded in
the thylakoid membrane
-protons are pumped into the thylakoid
space to form a proton gradient

24
 Light-Dependent Reactions
Photosystem I
-receives energy from an antenna complex
-energy is shuttled to P700 reaction center
-excited electron is transferred to a
membrane-bound electron carrier
-electrons are used to reduce NADP+ to
NADPH
-electrons lost from P700 are replaced from
the b6-f complex 25
 Light-Dependent Reactions
ATP is produced via chemiosmosis.
- ATP synthase is embedded in the
thylakoid membrane
-protons have accumulated in the thylakoid
space
-protons move into the stroma only through
ATP synthase
-ATP is produced from ADP + Pi
26
27
28
Noncyclic Photophosphorylation
• Photosystem II regains electrons by splitting
water, leaving O2 gas as a by-product
Primary
electron acceptor

Primary
electron acceptor

Photons

Energy for
synthesis of

PHOTOSYSTEM I

PHOTOSYSTEM II by chemiosmosis
How the Light Reactions Generate ATP and NADPH
Primary NADP
electron
acceptor
Energy
Primary to make 3
electron
acceptor 2

Light

Light

Primary
electron
acceptor

Reaction-
1 center NADPH-producing
chlorophyll photosystem

Water-splitting
photosystem
2 H + 1/2
31
 Carbon Fixation Reactions
To build carbohydrates, cells need:
1. energy
-ATP from light-dependent reactions

2. reduction potential
-NADPH from photosystem I

32
 Carbon Fixation Reactions
Calvin cycle
-biochemical pathway that allows for carbon
fixation
-occurs in the stroma
-uses ATP and NADPH as energy sources
-incorporates CO2 into organic molecules

33
 Carbon Fixation Reactions
carbon fixation – the incorporation of CO2
into organic molecules
-occurs in the first step of the Calvin cycle

ribulose-bis-phosphate + CO2 2(PGA)

5 carbons 1 carbon 3 carbons

The reaction is catalyzed by rubisco.

34
35
 Carbon Fixation Reactions
During the Calvin cycle, energy is needed.
The energy is supplied from:

- 18 ATP molecules
- 12 NADPH molecules

36
 Carbon Fixation Reactions
The energy cycle:

-photosynthesis uses the products of

respiration as starting substrates
-respiration uses the products of
photosynthesis as starting substrates

37
38
39
 Electron Transport Chain
The electron transport chain (ETC) is a
series of membrane-bound electron
carriers.
-embedded in the mitochondrial inner
membrane
-electrons from NADH and FADH2 are
transferred to complexes of the ETC
-each complex transfers the electrons to the
next complex in the chain
40
 Electron Transport Chain
As the electrons are transferred, some
electron energy is lost with each transfer.

This energy is used to pump protons (H+)

across the membrane from the matrix to
the inner membrane space.

A proton gradient is established.

41
 Electron Transport Chain
The higher negative charge in the matrix
attracts the protons (H+) back from the
intermembrane space to the matrix.

The accumulation of protons in the

intermembrane space drives protons into
the matrix via diffusion.

42
 Electron Transport Chain
Most protons move back to the matrix
through ATP synthase.

ATP synthase is a membrane-bound

enzyme that uses the energy of the proton
gradient to synthesize ATP from ADP + Pi.

43
 Oxidative
Phosphorylation

44
Electrons flow downhill
 Electrons move in steps from
carrier to carrier downhill to oxygen
 each carrier more electronegative
 controlled oxidation
 controlled release of energy

make ATP
instead of
fire!
46
• Glycolysis and thePhosphorylation
Oxidative citric acid cycle yield
NADH and FADH2.
• Both these electron carriers are energy-rich
molecules because their electrons have a
high transfer [redox] potentials.
• Oxidative phosphorylation is the process of
converting this high redox potential into
energy-rich ATP molecules.
• This process, together with the reactions
that form the electron carriers is often
called respiration.
47
ATP synthesis via oxidative phosphorylation
occurs via two separate systems in the
mitochondrion.
1. Electrons are “transported” via numerous
membrane-bound carriers from NADH to
O2. During these reactions, a proton
gradient is formed across the mitochondrial
inner membrane.
2. The proton-motive force in the gradient is
then harnessed to produce ATP.

48
Rather than occurring in a
single step, electrons
from NADH pass I
through groups of
carriers, mostly within
the mitochondrial inner II
membrane, eventually
reaching oxygen. III
The most interesting of
these carriers are three
groups of protein
complexes often
identified as IV
“Sites I, II, III, & IV.”
49
50
Electron Transport Chain
Sites I, II, III, and IV each contain numerous
protein subunits:

I
II
III

IV

52
 Oxidative Phosphorylation

• FADH2 delivers electrons to

the electron transport
pathway by reducing FADH2
ubiquinol (Q) to ubiquinone
(QH2).
• Recall that FADH2 is formed
in the TCA cycle when
succinate is oxidized to
malate. Electrons from
FADH2 reduce Q to QH2 and
flow through electron
transport, ending up on
oxygen.
• Consequently, less ATP is
formed from the oxidation of
53
FADH2 than from NADH.
The pathway of electron transport is easily determined by comparing reduction
potentials (E’0):

54
Diagram of electron transport chain and electron flow.

FADH2
FAD

55
(Garrett & Grisham, Biochemistry, 3rd ed., Brooks/Cole)
Another View of Electron Transport

56
(Mathews, et.al, Biochemistry, 3rd ed., Addison Wesley)
Complex I
Alternate Entries
Alternate Entries
Complex II
Complex III
Complex III
Complex IV
Complex IV
Electron Transport Chain
How is the proton-motive force created by
obligatory proton transport during passage
of electrons converted into high-energy
phosphate bonds in ATP?

65
Ephriam Racker (Cornell) discovered unique knob-like
structures on the matrix side of the inner membrane.
He removed these knobs with mild detergents and
mixed them with ATP. The ATP was immediately
hydrolyzed to ADP, so he named the knobs “ATPase.”

66
Protons flow through Oxidative Phosphorylation
the channel (F0) to the
large knob (F1)where
ATP is synthesized.
The “ATPase” knobs
described by Racker
destroy ATP by
converting it into ADP
+ Pi if they are not
attached to the F0
subunit.
When attached, they
catalyzed the opposite
reaction, namely ATP
synthesis. 67
 Structure of the ATP Synthase Complex
F1F0-ATP Synthase Subunits (E.coli)

Mass
Complex Subunit Number
(kD)

F0 a 1 30
b 2 17
c 9-12 8
F1 α 3 55
β 3 52
γ 1 30
δ 1 15
ε 1 5.6

68
• Paul Boyer finally put the puzzle together by
proposing that there must be three sites
with different binding affinities for the
substrate (ADP + Pi) and product (ATP).
• In fact, the three β-subunits interact in such
a way that when one assumes the β-empty
form, its neighbor to one side must assume
the β-ADP form, and the other neighbor the
β-ATP form.
• Thus, one complete rotation of the γ-subunit
causes each β-subunit to cycle through all
three of its possible confomations, and for
each rotation, three ATPs are synthesized
and release from the enzyme surface.
• Boyer received the Nobel Prize for this work
in 1997 (born and raised in Provo, Utah)
69
http://nobelprize.org/chemistry/laureates/1997/boyer-autobio.html
 ATP synthesis – byer’s “Binding-change Model”

Stryer, et.al, Biochemistry, 5th ed.

(Mathews, et.al, Biochemistry, 3rd ed., Addison Wesley)

70
(Garrett & Grisham, Biochemistry, 3rd ed., Brooks/Cole)
• ATP moves from the ATPmitochondrial
synthesis matrix to the
cytosol via a specialized membrane transport
protein, “ATP-ADP translocase.”
• Translocase is tightly coupled to the exchange of
ADP for ATP as ATP exits.

71
• Some NADH molecules are reduced in the
cytosol and must be transported into the
mitochonria for electrons to enter the
electron transport pathway.
• Two different “shuttles” are commonly
encountered:
– Glycerol 3-phosphate shuttle (transfers
electrons to FADH2 .
– Malate-aspartate shuttle (transfers electrons
to NADH) 72
• Malate-aspartate shuttle: (NADH 2e- NADH)

73
Oxidative Phosphorylation – Proton Flow

74
(Garrett & Grisham, Biochemistry, 3rd ed., Brooks/Cole)
• ATP synthesis can be
“uncoupled,” if the proton
gradient is prematurely
dissipated or impeded.
• Certain inhibitors of
electron transport act at
specific sites to stop
electron flow.
• Site I: amytal & rotenone
• Site III: antimycin A
• Site IV: CN-, N3-, CO

75
 Oxidative Phosphorylation (OP)
• Culmination* of energy-yielding metabolism in aerobic organisms

• Final stage of cellular respiration - Oxidative degradation of carbs, fats,

amino acids

• Energy of oxidation drives synthesis of ATP

• Photophosphorylation (PP): Means by which photosynthetic organisms

capture energy of sunlight

• Sunlight - ultimate source of energy in biosphere to make ATP

* the highest or climactic point of something, climax

• OP, PP - Accounts for most ATP synthesized by most organisms most of
the time
Subcellular localization

• OP in mitochondria, PP in chloroplasts

• ATP synthesis in mitochondria & chloroplasts based on chemiosmotic

theory

• Hypothesis by Peter Mitchell in 1961

• Transmembrane differences in proton concentration - reservoir for energy

extracted from biological oxidation reactions
Similarities between Oxidative phosphorylation & photophosphorylation

First: Both processes involve e- flow through a chain of membrane-bound

carriers (Figure 19-1)

Second: Free energy made available by this “downhill” (exergonic) e - flow is

coupled to “uphill” transport of H+ across a proton-impermeable membrane

• Free energy conserved as a transmembrane electrochemical potential

Third: Transmembrane flow of H+ down concentration gradient through

specific protein channels provides free energy

• Free energy used for ATP synthesis (ATP synthase)

 Mitochondria
• Metabolic role of mitochondria - critical to cellular/organism

• Defects in mitochondrial function - serious medical consequences

• Mitochondria - central to neuronal & muscular function

• Mitochondria - regulate whole-body energy metabolism

• Compromised mitochondrial function - Human neurodegenerative diseases,

cancer, diabetes, obesity

• Theory of aging - gradual loss of mitochondrial integrity

Mitochondrial functions:
1. ATP production
2. Thermogenesis (production of heat especially in body [as by oxidation])
3. Steroid synthesis
4. Apoptosis (programmed cell death)
• Eugene Kennedy & Albert Lehninger discovered (1948) - mitochondria
are site of oxidative phosphorylation

• Mitochondria has 2 membranes (Fig. 19-2a)

Outer membrane
• permeable to small molecules, ions - transmembrane channels (Porins :
integral membrane proteins family)

Inner membrane - selectively permeable

• Impermeable to most small molecules, ions, H+

• Transport possible through specific transporters

• Inner membrane - Respiratory chain, ATP synthase

Matrix contains
1. Pyruvate dehydrogenase complex
2. Citric acid cycle (CAC) enzymes
3. Fatty acid oxidation pathway enzymes
4. Amino acid oxidation enzymes & All fuel oxidation pathways (except glycolysis)

• Inner membrane segregates cytosolic & matrix intermediates & enzymes

• Specific transporters carry pyruvate, fatty acids, amino acids or their α-keto
derivatives into matrix for access to CAC machinery

• ADP, Pi transported into matrix

• Newly synthesized ATP transported out

• No. of Proteins in mammalian mitochondria - 1,100

• At least 300 proteins - unknown functions

FIGURE 19-2a Biochemical anatomy of a
mitochondrion

• Convolutions (cristae) inner membrane

provide a very large surface area

• Inner membrane of a single liver

mitochondrion may have more than
10,000 sets of electron-transfer systems
(respiratory chains) & ATP synthase
molecules, distributed over membrane
surface
• (b) Mitochondria of heart muscle, which have more profuse cristae &
thus a much larger area of inner membrane, contain more than three times
as many sets of electron-transfer systems as (c) liver mitochondria.
• Mitochondrial pool of coenzymes & intermediates is functionally separate
from cytosolic pool
• Mitochondria of invertebrates, plants, microbial eukaryotes are similar to
those shown here, but with much variation in size, shape, degree of
convolution of inner membrane
• Brain, skeletal & heart muscle, eye - high demand for aerobic metabolism

• These contain 100 -1000s mitochondria/cell

• In mitochondria of cells with high metabolic activity - >> densely packed,

cristae (Fig. 19–1b)

• Tissues with less-active metabolism (skin) << mitochondria fewer cristae

• During cell growth, division - mitochondria divide by fission* (like

bacteria)

• Sometimes, individual mitochondria fuse together

* Fission: action of dividing or splitting something into 2 or more parts

 Electrons Are Funneled to Universal Electron Acceptors
• OP: e- pass into respiratory chain

• Most e- arise by action of dehydrogenases

• e- funnel to universal electron acceptors

1. Nicotinamide nucleotides (NAD or NADP)
2. Flavin nucleotides (FMN or FAD)

1. Nicotinamide nucleotides-linked dehydrogenases

• Dehydrogenases act in catabolism – specific for NAD as e- acceptor (Table
19-1)

• Dehydrogenases present in - cytosol, mitochondria & others mitochondrial

& cytosolic isozymes

• NAD-linked dehydrogenases remove 2 H atoms from S

• 1 H transferred as a hydride ion (:H) to NAD (Fig.13-24)

• Other H is released in medium

• NADH, NADPH - water-soluble e- carriers

• NADH carries e- from catabolic reactions to their point of entry into ETC

• NADPH generally supplies e- to anabolic reactions

FIGURE 13-24 NAD and NADP.
(a) Nicotinamide adenine dinucleotide, NAD+, & its phosphorylated analog NADP+
undergo reduction to NADH & NADPH, accepting a hydride ion (two electrons and one
proton) from an oxidizable substrate. Hydride ion is added to either front (A side) or back
(B side) of planar nicotinamide ring.
(b) UV absorption spectra of NAD+ & NADH. Reduction of the nicotinamide ring
produces a new, broad absorption band with a maximum at 340 nm.
• Separate pools of NADPH, NADH in cell, with different redox potentials

• NADH, NADPH cannot cross inner mitochondrial membrane

• BUT their e- can be shuttled across

2. Flavoproteins

• Contain a very tightly, or covalently, bound flavin nucleotide (FMN, FAD)

(Fig. 13-27)

• Oxidized flavin nucleotide can accept either 1 e- (semiquinone form) or 2 e-

(FADH2 or FMNH2)

• e- transfer occurs as - flavoprotein has a higher RP (reduction potential)

than compound oxidized

• RP - quantitative measure of relative tendency of a given chemical species

to accept e- in an oxidation-reduction reaction
FIGURE 13-27 Oxidized, reduced FAD & FMN. FMN consists of structure
above dashed line on FAD (oxidized form). Flavin nucleotides accept 2 H atoms (2
e- & 2 H), both of which appear in flavin ring system. When FAD or FMN accepts
only 1 hydrogen atom, semiquinone, a stable free radical, forms.
• Standard reduction potential (SRP) of a flavin nucleotide (unlike NAD,
NADP)- depends on protein with which it is associated

• Interactions of protein functional groups - distort e- orbitals in flavin ring

• It changes stabilities of oxid. & red. Forms

• Relevant SRP is that of particular flavoprotein

• Relevant SRP is not of isolated FAD or FMN

• Flavin nucleotide should be considered part of flavoprotein’s active site

rather than a reactant or product in e- transfer reaction

• As flavoproteins can serve as intermediates:

1. When 2 e- donated, e.g. dehydrogenations
2. When 1 e- accepted, e.g. reduction of a quinone to a hydroquinone
Fig.19-30 Glycerol 3-phosphate shuttle. This alternative means of moving reducing equivalents
from cytosol to mitochondrial matrix operates in skeletal muscle & brain. In cytosol,
dihydroxyacetone phosphate accepts 2 reducing equivalents from NADH in a reaction catalyzed by
cytosolic glycerol 3-phosphate dehydrogenase.
An isozyme of glycerol 3-phosphate dehydrogenase bound to outer face of inner membrane then
transfers 2 reducing equivalents from glycerol 3-phosphate in intermembrane space to ubiquinone.
Note that this shuttle does not involve membrane transport systems
Fig. 19-29 Malate-aspartate shuttle. Shuttle for transporting reducing
equivalents from cytosolic NADH into mitochondrial matrix is used in
liver, kidney, & heart.

1 NADH in cytosol (intermembrane space) passes 2 reducing

equivalents to oxaloacetate, producing malate. 2 Malate crosses inner
membrane via malate–α-ketoglutarate transporter. 3 In matrix, malate
passes 2 reducing equivalents to NAD+, & resulting NADH is oxidized
by respiratory chain; oxaloacetate formed from malate cannot pass
directly into cytosol. 4 Oxaloacetate is first transaminated to aspartate,
& 5 aspartate can leave via glutamate-aspartate transporter. 6
Oxaloacetate is regenerated in cytosol, completing cycle.