III. Metabolism Photosynthesispeople.uleth.ca/~steven.mosimann/bchm3300/Bchm3300_L10.pdf ·...

Biochemistry 3300 – Photosynthesis I Slide 1

III. Metabolism

Photosynthesis

Department of Chemistry and BiochemistryUniversity of Lethbridge

Biochemistry 3300


Overall Scheme of Photosynthesis

Photosynthesis can be separated into“light” and “dark” reactions:

CO2 + H2O → O2 + (CH2O)

Yield Energy

ConsumeEnergy

Dark reaction (consume energy):

2H2O + 2NADP+ → 2NADPH + 2H+ + O2

Light reaction (yield energy):

Light Reactions

Dark Reactions


Photosynthesis Takes Place in Chloroplasts

Stroma contains flattenedvesicles (thylakoids), stackedin grana.→ contains membrane-associated

ATP synthesis machinery

Light reactions: thylakoid membranes

Dark reactions: stroma

They have two membranes:→ outer membrane is porous

the inner is not.


Electromagnetic Radiation

The Spectrum of electromagnetic radiation, and the energy of photons in thevisible range of the spectrum.

Note: 1 mole of photons = einstein


Determination of the Action Spectrum of Photosynthesis

Oxygen electrode allows measurement of O2 production.Action spectrum describes the relative rate of photosynthesis forillumination with a constant number of photons of different wavelengths.→ correlates with absorption spectra of photosynthetic chromophores


Absorption of Visible Light by Chromophores.

A combination of chlorophylls together with accessory pigments allow most of the sun’s energy to be harvested.

Absorption in the red and blueareas by plants (chlorophylls)gives them their green color.→ only green light is left to

reflect or transmit

Absorption of 1 mol of photons yields 170-300 kJ


Important Chromophores in Photosynthesis

Primary pigments are Chlorophylls a, b (plants), Bacteriochlorophyll (bacteria),and phycobilins (algae, cyanobacteria)



Primary pigments are Chlorophylls a, b (plants), Bacteriochlorophyll (bacteria),and phycobilins (algae, cyanobacteria)


PhycobilisomesLight Harvesting Complex of cyanobacteria

Phycobilins are covalently linkedto specific binding proteins

→ phycobiliproteins

Phycobiliproteins associate in highly ordered complexes.

→ phycobilisomes→ light-harvesting structure in e.g. red algae

Phycobilin pigments bound to specific proteins form complexes calledphycoerythrin (PE), phycocyanin (PC), and allophycocyanin (AP)



β-Carotene (a Carotenoid) and Lutein (a Xanthophyll) are accessory pigmentsin plants.


The Light-harvesting Complex from Plants

The functional complexis a trimer of LHCII.

→ containing 6 Lutein and36 Chlorophyllmolecules

7 Chlorophyll a5 Chlorophyll b2 Lutein

View along the membrane plane


Light HarvestingComplex (non-plant)

Green – chlorophyllYellow – accessory pigments


Photosystems and the Thylakoid Membrane.

Thylakoid membranes arefunctionally arranged into photosystems with 50-200chromophores per system

Light is transduced to chemicalenergy only in the photochemicalreaction center.→ a special pair of chlorophyll a

molecules

The function of most of thechromophores in a photosystem isto harvest light photons and totransfer, by excitation transfer,the energy successively to thereaction center.


Exciton and Electron Transfer


Functional Modules of the Photosynthetic Machinery

There are several classes of photosynthetic reaction centers

I. Single reaction centers → Type I Fe-S and → Type II Pheophytin

-Quinone

II. Double reaction centers (Cyanobacteria, algae, and vascular plants)



Purple bacteria – Light photons excite the P870 chromophore

→ Electron flow is cyclic

→ Proton gradient is generated

→ ATP synthesis


Photoreaction Center of Purple Bacteria

PDBid 1PRC

X-ray structure of purple bacterium reaction center.

→ 3 integral membrane subunits→ Cytochrome c outside the membrans

Space-filling models are thechromophores in the membrane andthe cytochrome c part.


The Electron Transfer

The Structure together with rapid reaction kineticsallowed the assignment of the sequence of eventsin electron transfer.

1. Lights excites the Chl2 pair2. e- pass to Pheophytin3. e- move to Quinone A4. e- flow through non heme iron

to Quinone B (diffusible)

5. The electron hole in Chl2 is filled by e- transfer a heme of Cyt c

6. Fully reduced Quinone B diffuses to Cyt bc1 and initiate the formation of the proton gradient

very slow!



→ Electron flow is cyclic

→ Proton gradient is generated

→ ATP synthesis

Green-sulfur bacteria also have a P840 and Cytochrome bc1

Coupled reaction:H2S is oxidized to S, then to SO4

2- toprovide electrons to P840

But some of the e- are passed to theredox protein ferredoxin, through a reductaseto produce NADH

That’s a problem! What could be the solution?


Two Reaction Centers Act in Plants

In plant chloroplasts photosynthesisrequires two reaction centers.

The “Z Scheme” is not cyclic.

H2O is oxidized and NADP+ isreduced to NADPH.

Two photon absorptions are neededto drive the reduction in PSI/PSII

Oxygen is evolved in a specificcomplex (water-splitting complex)

A proton gradient is developed



PS II resembles the purple bacteriareaction center.PSI resembles the green-sulfurbacteria center.

H2O oxidation in plants is analogous to the H2S oxidation by green-sulfur bacteria.→ activated e- are replaced→ oxygenic photosynthesis

Plastocyanin is a diffusible smallprotein (one e- carrier) thatshuttles between PSII and PSI

All O2 evolving organisms havetwo coupled photosystems.



Overall Z-scheme

2 H2O + 2 NADP+ + 8 hv → O2 + 2 NADPH + 2 H+

One electron transferred per 2 photons absorbed, forming O2 requires4 electron transfers (8 photons)


PSII Mechanism

Transmembrane protein (D1/D2 dimer), but electrons flow only through D1.

e- flow from the Mn4

center (active site ofthe water-splitting enzyme) to replacee- excited by lightin P680.

After excitations, e-

flow through carriersto quinone PQB and tocytochrome b6f


PSI Mechanism

Excited e- from P700 [(Chl)2pair] is transferred through ferredoxin.→ Ferredoxin:NADP+ oxidoreductase transfers e- to NADP+

→ NADPH and ferredoxin(ox)

Plastocyanin (soluble Cu-containig transporter) resupplies P700+ with e-


The PSI Complex is Associated with Antenna Chlorophylls

PDBid 1JBO

The proteins in the complex hold the components rigidly in orientations thatmaximize efficient exciton transfer betweenexcited antenna molecules towards P700

Trimeric structure viewed from thethylakoid lumen.

PSI monomer with proteins removed

Fe-S centers


Cytochrome b6f Complex Links to Photosystems I and II

Connection between PSII and PSI centers→ Plastoquinol formed in PSII is oxidized

by the cytochrome b6f complex→ a Q-cycle generates a proton gradient→ H+ are pumped from stroma to lumen

PDBid 1VF5


Proton and Electron Circuits in Thylakoids

An ATP synthase in Chloroplasts harnesses theproton gradient generated by the cytochrome b6fcomplex to synthesize ATP.


Water-splitting Activity in the Oxygen-evolving Complex

The ultimate source of e- is Water → splitting requires energy of 4 photons.

Each photon into P680 ejects an e- → replaced by e- from Mn complex

The Mn4+ complex accepts four e- from 2 H2O → Oxygen (O2)


Alternatives

An alternative path of light-inducedelectron flow allows variation inthe amounts of ATP and NADPHthat are made.

→ Cyclic pathway e- partitioned at Ferrodoxin some return to Cyt b6f

→ only PSI is involved in this pathway→ resembles the green-sulfur bacteria

pathway

→ No O2 or NADPH is made, butH+ is pumped and ATP is synthesized.


Cyanobacteria

Cyoanobacteria use cyt b6f, cyt c6

and plastoquinone for both

- oxidative phosphorylation→e- flow from NADH to O2

- photophosphorylation →e- flow from O2 to NADPH

→ occure at the same membrane(no motiochondria or chloroplasts exist)

Both processes are accompanied byH+ movement across the membrane


ATP Synthase of Chloroplasts is similarto Mitochondrial Enzyme

ATP is synthesized in chloroplasts by a large complex with thetwo functional components CFO and CF1

CFO is a transmembrane proton pore andCF1 is a peripheral membrane protein similarin structure and function to mitochondrial F1

ADP + Pi → ATP + H2Olight


A Single Protein Absorbs Light in Halophilic Bacteria

PDBid 1C8R

Bacteriorhodopsin has seven membran-spanning α-helices. The chromophore all-trans retinal is covalently attached

via a Schiff base to an ε-amino groupof Lysine.

A series of Asp and Glutogether with closeassociated H2O providea transmembrane pathfor protons.


Light-driven Proton Pumping in Bacteriorhodopsin

In the dark the Schiff base is protonated.→ Illumination photoisomerizes the all trans-retinal → H+ is transferred to Asp85


Light-driven Proton Pumping in Bacteriorhodopsin

The protein’s principle H+

pumping motions are remarkebly small→ groups move about ~1Å

These causes pKa changes in various residues.→ H+ transfer

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