Structure and function of Cytochrome b6f complex and Photosystem-I

Under the Guidance of

Prof. S. M. Prasad Presented By Rajnish Kumar M.Sc-3rd Sem.(Botany)

Department of botany

Photosynthesis

Photosynthesis is a biochemical reaction which convert solar energy to chemical energy.

Photosynthesis is anabolic process.

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

In photosynthesis, plants are taken CO2 from atmosphere and H2O from soil to formed carbohydrate and Oxygen. In the presence of sunlight and chlorophyll.

Sun lightChlorophyll

Photosynthesis in higher plants

In plants light energy convert into chemical energy or photosynthesis are takes place in chloroplasts.

In higher plants the most active photo -synthetic tissue is mesophyll of leaves.

Mesophyll cells have many chloroplast, which contain the specialized light-absorbing green pigments. i.e.-chlorophyll.

In photosynthesis plant use solar energy to oxidize water, thereby releasing oxygen, and to reduce carbon dioxide, thereby forming large carbon compounds, primarily sugars.

The location and structure of - chloroplasts

LEAF CROSS SECTION MESOPHYLL CELL

LEAF

Chloroplast

Mesophyll

CHLOROPLASTIntermembrane space

Outermembrane

Innermembrane

ThylakoidcompartmentThylakoidStroma

GranumStromaGrana

Structure of Chloroplast Chloroplast has two membranes-

1-outer membrane – permeable. 2-inner membrane - enclosed inner compartment.

Each membrane is made up of lipoproteins.

The width of each membrane is 50Å.

Inner compartment- This membrane enclosed sacs called

Thylakoids involving paired folds (lamellae) by stacking form "Grana".

The soluble portion of the chloroplast is the "Stroma".

The aqueous compartment (Stroma) contained several enzymes, So it is the site of carbon fixation (synthesis of carbohydrate - Dark reaction).

The interior of the thylakoid vesicles is the "Thylakoid space" or "Thylakoid lumen".

Thylakoid membrane contains Pigments like chlorophyll and carotenoids, for Photophosphorelation and the enzymes for ATP synthesis.

Chlorophylls - primary light-absorbing pigments. Carotenoids - accessory pigments.

Ultrastructure of chloroplast

CHLOROPLAST PIGMENTS A Chloroplast pigment (accessory pigment;photosynthetic pigment; antenna pigment) is a pigment that is present in chloroplasts or photosynthetic bacteria and captures the

light energy necessary for photosynthesis. Chloroplasts contain several pigments - Chlorophyll a - Blue-green pigment.

Chlorophyll b - Yellow-green pigment. Carotenoids - An orange pigment. Xanthophyll - An Yellow pigment.

Chlorophyll Chlorophyll is a light absorbing green

pigment. Chlorophyll is a lipid soluble pigment. Chlorophyll contains a polycyclic, planar

tetrapyrrole ring structure.

The Central metal ion in chlorophyll is Mg2+ .

Chlorophyll has a cyclopentanone ring (ring-V) fused to pyrrole ring III.

Chlorophyll has a long Phytol chain on Pyrrole ring-IV

Chl-a & chl-b are mostly found in green plants.

In Chlorophyll-a (C55H72O5N4Mg)-present CH3 Group on II Pyrrole ring.

In Chlorophyll-b (C55H70O6N4Mg)-CH3 Group is replaced by CHO Group.

Carotenoids are linear molecules with multiple conjugated double bonds.

Carotenoids are lipid soluble pigment.

Carotenoids are Orange pigment (400-500 nm).

Carotenoids are accessory pigment.

Carotenoids are linear polyenes that serve as both antenna pigments and photo protective agents.

Carotenoids

Nature of lightSunlight is a type of energy, which are

called as radiant energy or electromagnetic energy.

Light has characteristics of both a wave and particle.

A wave is characterized by a wavelength.

The distance between two successive wave crests is known as wavelength.

Wavelength is denoted by the Greek letter lambda(.

Wavelength is measure in nanometer(nm).

Frequency is the number of wave crests that pass an observer in a given time.

Frequency is denoted by Greek letter nu().

Frequency is measure in Hertz(Hz).

The relationship between wavelength and frequency is-

Where, C = Speed of light.(3.8×10 8 ms -1) = Wavelength. = Frequency.

c =

Light is also travels a particles. These particles or packets are called Photon.

Each photon contains an amount of energy that is called a Quantum(pl.-Quanta).

The energy (E) of photon depends on the frequency of the light according to a relation known as Planks law-

Where, E = Energy. H = Planks constant(6.626×10-34 js). .

E = h

When sunlight is pass through a prism then will we found found different colour of rays which are called as visible spectrum.

The complete visible spectrum is made up of seven colour. Which are found in form of VIBGYOR .

ROLE OF PHOTOSYNTHETIC -PIGMENTS IN PHOTOSYNTHESIS

When sunlight is falls on leaves. In which 83% is absorbed by leaves, 12% is reflected & 5% is transmitted.

In photosynthesis only 4% sunlight is used by chlorophyll and 79 % sunlight is diffused in atmosphere in the form of heat.

The light absorbing complex is called as photosynthetic pigment. for example-

Chl-a, Chl-b, Carotenoids etc.

These photosynthetic pigment are found in membrane of thylakoid and mainly absorbed Blue, Violet, Red & Orange rays.

Chlorophyll-b, carotenoids & some kinds of Chl-a are play the important role in Antenna molecule.

These antenna molecule absorb light energy and transfer to reaction center.

After absorb the light energy pigment molecules are excited or unstable.

chl. + h chl.* Excited chlorophyll can re-emit a photon in

the form of heat and thereby returns to its ground state. These process is known as fluorescence.

Mechanism of Photosynthesis - Source of Librated Oxygen In Photosynthesis

Before 1930, Scientists consider, the one molecule of CO2 and one molecule of H2O are formed one molecule of formaldehyde in the presence of sunlight.

These formaldehyde give rise to glucose molecule after Polymerization. CO2 + H2O CH2O

+ O2

6CH2O C6H12O6

Sunlight Polymerization

Formaldehyde

Hexose sugar

But formaldehyde are poisonous compound, which causes death of plants. Hence in photosynthesis the formation of formaldehyde is not possible.

In 1930, Cornelius B. Van Niel proved that sulfur bacteria are formed carbohydrate from H2S and CO2 in the presence of light and produced sulfur. 6CO2 +12H2S C6H12O6 +

6H20 +12 S Light

Hence Van Niel suggest that sulfur are separated by synthesis of sulfur bacteria. therefore in photosynthesis oxygen is separated by decomposition of water.

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

The above hypothesis was proved by Ruben in 1941 on his experiment by using O18 atomic weight of isotopes and he concluded oxygen are come out from water molecule in photosynthesis.

So the complete equation of photosynthesis is written fallowing-

LightChlorophyll

Oxygenic and An-oxygenic photosynthesis

Oxygenic photosynthesis produce oxygen and it is found in green plants and cyanobacteria.

CO2 + H2O SUGAR + OXYGEN

An-oxygenic photosynthesis does not produce oxygen and it is found in green and purple photosynthetic bacteria.

CO2 + H2S SUGAR + SULPHUR

Modern concept of mechanism of photosynthesisPhotosynthesis is a oxidation-reduction

process. In which water oxidize, thereby releasing oxygen, and carbon dioxide reduce, thereby forming large carbon compounds, primarily sugars.

Photosynthesis are completed in two phases - 1. Photocemical reaction / light dependent

reaction.2. Chemical dark reaction / light

independent reaction.

Light dependent reaction It is occurs in grana of chloroplast.

In the chloroplast, light energy is converted into chemical energy by two different functional units called photosystems.

Photosystem are two type -1. Photosystem-I / Photo-act-I 2. Photosystem-II / Photo-act-II

Each photosystem has over 200 molecules of chlorophylls and about 50 molecules of carotenoids.

LIGHT HARVESTING COMPLEXES (LHCs) The light

harvesting complex (antenna complex) is an complex of protein and chlorophyll molecules embedded in the thylakoid membrane of plants, which transfer light energy to one chlorophyll a molecule at the reaction center of photosystem.

Those LHC is associated with PS-I is called LHC-I protein.

Those LHC is associated with PS-II is called LHC-II protein.

DISTRIBUTION OF PHOTOSYNTHETIC PROTEINS

The thylakoid membrane of chloroplast are found in two forms stacked (appressed) and un-stacked (non-appressed) region.

PS-II are found in mainly stacked region.

PS-I & ATP Synthase are found in mainly un-stacked region. which are attached to stroma.

Cyt-b6-f complex are similarily found in both stacked & un-stacked region.

Cytochrome b6f Complex The cytochrome b6f  complex  is an enzyme

found in the thylakoid membrane in chloroplasts of plants, cyanobacteria, and green algae, that catalyze the transfer of electrons from plastoquinol to plastocyanin.

During photosynthesis, the cytochrome b6f complex transfers electrons from Photosystem II to Photosystem I, whereby pumping protons into the thylakoid space and creating an electrochemical (energy) gradient where it is later used to create adenosine triphosphate (ATP).

The cytochrome b6f complex is a large multi -subunit protein with Seven prosthetic groups.

The cytochrome b6f complex is a dimer, with each monomer composed of eight subunits.

The cytochrome b6f complex is consist of four large subunits-

STRUCTURE OF CYTOCHROME b6f COMPLEX

cytochrome f (PetA ) with a c-type cytochrome,

cytochrome b6 (PetB) with a low- and high-potential heme group,

Rieske iron-sulfur protein (PetC) containing a [2Fe-2S] cluster, and

subunit IV(PetD); along with four small subunits : PetG, PetL, PetM, and PetN.

H2O →photosystem

II→ QH2 → Cyt b6f → Pc → photos

ystem I → NADPH (1)

QH2 → Cyt b6f → Pc → photosystem I → Q (2)

Reaction mechanismThe cytochrome b6f  complex is responsible for "non- cyclic" (1) and "cyclic" (2) electron transfer between two mobile redox carriers, plastoquinone (QH2) and plastocyanin (PC).

Cytochrome b6f catalyzes the transfer of electrons from plastoquinol to plastocyanin, while pumping two protons from the stroma into the thylakoid lumen:QH2 + 2Pc(Cu2+) + 2H+ (stroma) → Q + 2Pc(Cu+) + 4H+ (lumen)

This reaction occurs through the Q cycle as in Complex III. Plastoquinone acts as the electron carrier, transferring its two electrons to high- and low-potential electron transport chains (ETC) via a mechanism called electron bifurcation.

Q - Cycle

(PQH₂) is oxidised & one of 2 e⁻ is passed a linear e⁻ transport chain toward PS-I & other e⁻ goes through a cyclic process that increases the number of protons pumped across the membrane.

(A) The non-cyclic or linear processPQH₂ Produced by PS-II is oxidised near the lumenal side of the complex, transfering its 1e⁻ to Rieske Fe-S protein & 1e⁻ to b-type Cyt. & expelling 2 protons to lumen. The e⁻ transferred to cyt f & to PC which reduces P700 of PS-I. The reduced b-type cyt transfers e⁻ to other b-type cyt which reduces PQ to plastosemiquinone (PQ⁻) state.

(B) The cyclic process A second PQH₂ is oxidised , with 1 e⁻

going from FeSR to pc & finally to P700. The 2nd e⁻ goes through 2 b-type cyt & reduces the plastohydroquinone, at the same time picking up two protons from stroma ,Overall, 4 protons are transported across the membrane for every 2 e⁻ delivered to P700.

Biological function In photosynthesis, the cytochrome b6f

complex functions to mediate the transfer of electrons between the two photosynthetic reaction center complexes, from Photosystem II to Photosystem I, while transferring protons from the chloroplast stroma across the thylakoid membrane into the lumen.

Electron transport via cytochrome b6f is responsible for creating the proton gradient that drives the synthesis of ATP in chloroplasts.

In a separate reaction, the cytochrome b6f complex plays a central role in cyclic photophosphorylation, when NADP+ is not available to accept electrons from reduced ferredoxin.

This cycle results in the creation of a proton gradient by cytochrome b6f, which can be used to drive ATP synthesis.

It has also been shown that this cycle is essential for photosynthesis in which it is proposed to help maintain the proper ratio of ATP/NADPH production for carbon fixation.

PHOTOSYSTEM-I (PS-I)Photochemical events are similar to those in PS-II.

PS-I is composed of a heterodimer of proteins that act as ligands for most of the electron carriers.

light is absorbed by antenna molecules and the energy is transferred to P700 (reaction center) by resonance energy transfer.

Chlorophyll-a & Chlorophyll-b The PS-I reaction center and its associated

antenna pigments and electron transfer proteins, as well as the coupling-factor enzyme that catalyzes the formation of ATP, are found almost exclusively in the Stroma lamellae and at the edges of the grana lamellae.

PHOTOSYSTEM-I• Multi-subunit protein complex.

• Contains about 14 different proteins:

a)Core proteins –PsaA , PsaB.

b)Peripheral proteins on stromatal side-PsaC, PsaD, PsaE ,

c)Integral membrane proteins-PsaF, PsaG, PsaH, PsaI, PsaJ, PsaK, PsaL, PsaM.

d)Lumenal protein- PsaN.

Approx. 100 chlorophyll molecules.

12-16 carotenes.

Phylloquinone.

Additional chain of 5 electrons acceptors:A0, A1and 3(4fe-4s) centers :fx, FA, FB.

The excited reaction center P700* loses an electron to an electron acceptor, A0 (like pheophytin in PS-II) creating A0

- and P700+.

This results in charge separation at the photochemical reaction center. P700+ is a strong oxidizing agent ,

It acquires an electron from plastocyanin, a soluble cu-containing electron transfer protein.

A0- is a strong reducing agent. It passes

its electrons through a chain of carriers leading to NADP+.

ELECTRON TRANSFER PATHWAY IN PS-I

A0 passes its electrons to Phylloquinone(A1)

A1 passes it to an fe-s protein.

Fe-s protein passes the electron to ferredoxin(fd) (another fe-s protein).

The electron is then transferred to a ferredoxin NADP reductase. (flavoprotein) The electron is transferred from reduced fd to NADP+.

USE OF PHOTOSYSTEM - I

Photosystem-I used in cyclic and noncyclic photophosphorylation.

While photosystem-II used in only noncyclic photophosphorylation.

Cyclic Photophosphorylation In cyclic electron transfer, the electron

begins in a pigment complex called Photosystem-I.

Then passes from the primary acceptor to ferredoxin, then to cytochrome b6f and then to plastocyanin before returning to chlorophyll.

This transport chain produces a proton-motive force, pumping H+ ions across the membrane; this produces a concentration gradient that can be used to power ATP synthase during chemiosmosis.

This pathway is known as cyclic photophosphorylation, and it produces neither O2 nor NADPH.

In bacterial photosynthesis, a single photosystem is used, and therefore is involved in cyclic photophosphorylation. It is favored in anaerobic conditions and conditions of high irradiance and CO2 compensation points.

In noncyclic photophosphorylation, is a two-stage process involving two different chlorophyll photosystems. Being a light reaction, non-cyclic photophosphorylation occurs in the frets or stroma lamellae.

First, a water molecule is broken down into 2H+ + 1/2 O2 + 2e− by a process called photolysis (or water-splitting). The two electrons from the water molecule are kept in photosystem II, while the 2H+ and 1/2O2 are left out for further use.

Then a photon is absorbed by chlorophyll pigments surrounding the reaction core center of the photosystem.

Noncyclic Photophosphorylation

The light excites the electrons of each pigment, causing a chain reaction that eventually transfers energy to the core of photosystem II, exciting the two electrons that are transferred to the primary electron acceptor, pheophytin.

The deficit of electrons is replenished by taking electrons from another molecule of water.

The electrons transfer from pheophytin to plastoquinone, which takes the 2e− from Pheophytin, and two H+ atoms from the stroma and forms PQH2, which later is broken into PQ, the 2e− is released to Cytochrome b6f complex and the two H+ ions are released into thylakoid lumen.

The electrons then pass through the Cyt

b6and Cyt f. Then they are passed to plastocyanin, providing the energy for hydrogen ions (H+) to be pumped into the thylakoid space.

This creates a gradient, making H+ ions flow back into the stroma of the chloroplast, providing the energy for the regeneration of ATP.

The photosystem -II complex replaced its lost electrons from an external source; however, the two other electrons are not returned to photosystem -II as they would in the analogous cyclic pathway.

Instead, the still-excited electrons are transferred to a photosystem I complex, which boosts their energy level to a higher level using a second solar photon.

The highly excited electrons are transferred to the acceptor molecule, but this time are passed on to an enzyme called Ferredoxin-NADP+ reductase which uses them to catalyse the reaction (as shown):

NADP+ + 2H+ + 2e− → NADPH + H+

This consumes the H+ ions produced by the splitting of water, leading to a net production of ½ O2, ATP, and NADPH+H+ with the consumption of solar photons and water.

Effect of herbicides or weedicides on ETC

Herbicides are classified into three categories-

1. Urea Herbicides-Monuron And Diuron(DCMU)

2. Triazine Herbicides-Atrazine And Simazine.

3. Bypyridilium Herbicides-Diquat And Paraquat.

Urea and Triazine herbicides are absorbed in plants through root and then transfer to leaves. and it block the electron flow in plastoquinone. Therefore the electron transport will be stopped in PS-II to PS-I.

Bypyridilium herbicides are also known as Viologen dyes. It blocks the electron flow in Fe-S to Fd in PS-I. These herbicides are taken electron from PS-I and performed reaction with O2 and give rise to O-

2 (super-oxide ion).which are harmful to chloroplast component specially lipids. Therefore it is also harmful to chlorophyll.

Fe-S Fd

Plant Physiology - Taiz & Zeiger.Plant Physiology - B. Salisburg & C. W. Ross.

Life Science F & P-I: Pranav Kumar & Usha -

Mina.Wikipedia, the Free Encyclopedia.

REFERENCES

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