Lectures 10 and 11Introduction to Plant Physiology and Primary Metabolism

•Reading for Lectures 10 and 11Chapter 10 in Plants, Genes and Crop Biotechnology, p.240-269 (on Reserve in the library)---------------------------------------------------------------------------------Lecture 10: “Getting and Using Carbon for Energy and to BuildPlant Molecules”

-Genes encode enzymes. -Enzymes organize chemistry.-Enzymes build simple molecules (nucleotides, amino acids,sugars, etc.).-Other enzymes use these small molecules as subunits to build larger molecules (fatty acid membranes, DNA/RNA, proteins, cell walls, etc.)

Subunits Make Longer ChainsFig. 1 - nucleotides = DNA

Slide 10.1

Glucose, Fructose, Sucrose, Starch cellulose

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Fatty acids- 2C Acetyl CoA for compartment membranes:

Slide 10.2

Figures from :Plants, Genes and Agriculture,pp.86 and 89M. Chrispeels and D.SadavaJones and Bartlett Publishers, Boston, 1994

How does the cell make these subunits?

•Enzymes are creating C-C, C-P, C-N, C-O, O-P, C-S, etc. bonds.•Predominantly, the backbone consists of C-C bonds.•These subunits are constructed using 2Carbon to 6Carbon molecules as the backbone (to which N, O, P, S are added).

This lecture will answer the following 4 questions:1. Where does the carbon come from? _________2. To form C-C (and C-O, O-P, C-S, C-N, etc) bonds, energy is required. Where does this energy come from? _________How?3. How are the 2C-6C subunits formed from CO2?4. How are the 2C-6C subunits stored for future needs of the plant?

1. Fundamentally, why does life need energy?Negatively-charged electrons are attracted to the positively-charged nucleus (protons) of the same atom.

The electron can also be attracted to the nucleus of another atom.For example, an electron which is far away from its own nucleus(and less susceptible to positive forces) can more easily be attracted to the nucleus of another atom. Thus, all else being equal , it would require less energy to pull an electron from an outer location towards another nucleus, thanan electron in an inner orbiting location (orbital).

What is a chemical bond?

Bonds break and form because electron-sharing between atoms ormolecules begins or ends (as the electrons move further away orcloser to each nucleus).

Loss of an electron away from a nucleus = __________Gain of an electron closer to a nucleus = “___________

Similarly, to pull an electron further away from its nucleus requires energy to overcome forces of positive attraction. Theseelectrons possess more energy than electrons held in a moreStable conformation, such as closer to the nucleus.

Slide 10.3

Photosynthesis -- the energy from sunlight is absorbed by chlorophyll in close proximity to 2H20. An electron ofchlorophyll is excited (becomes in a high energy state), is pulledaway from its nucleus and transferred to an adjacentmolecules.

Chlorophyll then constantly replaces these “lost”electrons pulling them away from the nuclei of 2H20 (thus liberating O2 and 4H+ protons).

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2. Where do plants get energy from?

Slide 10.4

Chlorophyll is a large, bulky molecule, which is not diffusible and its energy cannot be used to build/break other bonds in the cell (for example to build DNA). Chlorophyll is embedded in membranes inside the chloroplast. Why is this important?

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The “lost” electrons from chlorophyll are passed onto nearbymolecules also embedded in the chloroplast membrane. TheseMolecules can absorb and give away excess electrons. Why are electrons passed along a chain like this?

Slide 10.5

3. Electron Transfer in the Chloroplast Generates CofactorsAt each electron transfer step in the chloroplast, the high-energy electrons go into a more stable orbital around the nucleus of the electron-accepting molecule.

As a result, energy is released, but gradually. A sudden drop in the energy state of an electron would releasemost of the energy as heat (not useable energy).

Slide 10.6

3. High Energy CofactorsThe electron energy gradually lost while exchanged betweenreduction-oxidation (redox chain) molecules in the chloroplast isused to boost the energy of electrons of diffusible molecules (energy cofactors).

These cofactors aid in pulling and pushing electrons between the nuclei of any two reactant molecules in the cell allowing themto form bonds such as C-C bonds. How?

In the cofactors, the high-energy electrons are unstable and attracted to the nuclei of other molecules. As the electrons leave the nucleus of the cofactor atom, they release the energy which kept the electrons in orbital; this energy can be used to pull/push electrons from other molecules to/from one another, to overcome energy activation barriers. This permits the breaking and forming of new bonds between reactant molecules. To do this, the cofactors temporarily bind to reactant molecules, and pull and push their electrons to overcome activation barriers; these reactant electrons can then participate in new bonds between two reactant molecule nuclei.

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NAD(P)H ATP

Slide 10.7

From Biochemistry and Molecular Biology of Plants(W.Gruissem, B. Buchanan and R.Jones p.679 and p.834

ASPP, Rockville MD, 2000

The low-energy (oxidized) form of the cofactors are _________, while the high-energy, high-electron-containing "reduced" forms are __________.

Therefore, the purpose of photosynthesis is to produce high-energy cofactors. Photosynthesis does NOT create C-C bonds.

Why can't chlorophyll or associated electron-acceptor molecules directly be used as a cofactor?

The focus of the next step is to use this energy to form C-Cbonds. Why?

C-C bonds must be formed to create the backbone of the cell's molecules (such as fats, cellulose). There is a second reason to form C-C bonds however: The cell does not store abundant amounts of ATP/NADPH cofactors, because they are very reactant. Instead, evolution chose to use the electrons held between the nuclei of carbon atoms (C-C bonds) as the storage form for energy. C-C molecules are not reactive and very stable (eg. fats, oils, carbohydrates).

Where does this Carbon come from and how is the C-C bondFormed and stored?

Slide 10.8

5. “Fixing” (Reducing) CarbonThe carbon is derived from CO2. To form C-C bonds, it would have been rational for evolution to take C from 2 CO2 atoms and join them together.

Apparently, evolution could not create an enzyme to do this. Instead, the only known enzyme that can take C from CO2 attaches it to a 5C acceptor, to create an unstable 6C molecule, which then breaks down into 2 x 3C molecules.

This enzyme is called RuBisCo (Ribulose BisphosphateCarboxylase). The addition of C to 5C requires ATP + NADPH.

This "carbon-fixation" reaction has three major problems associated with them. What are these and what are the consequences for agriculture?1a.1b.

2.

3. Slide 10.9

6. Biochemistry Uses CyclesThe chloroplast must constantly generate 5C acceptor molecule. How was this problem solved?

To go from 3C back to the 5C acceptor molecule, 9 more enzymes are involved. Along with Rubisco and 2 other enzymes, these form the Calvin Cycle.

Therefore, the majority of the enzymes of the Calvin Cycle have nothing to do with removing C from CO2 to form C-C bonds, but rather, they are there to generate the 5C acceptor molecule!! Is there a more efficient way to do this? Evolution didn't create one, but can humans?!!

Slide 10.10

7. What are the Critical Factors Regulating Rates of PSS/Carbon Fixation via the Calvin Cycle in Crops?

CO2/O2 enter the chloroplast via stomates in leaves-increase in CO2 enhances rate of carbon fixation. -atmospheric CO2 is 0.03%. Commercial greenhouses increase this to0.08% to increase yields of vegetables and ornamentals. Up to 0.1% isvery efficient under optimal light, temp, water and Nitrogen

What will be the effect of global warming and enhanced CO2???

-stomates open during day/closed at night. Enhanced H20 allows stomates to stay more open, allowing more CO2 to flow in.

-problem: C-fixing enzyme (Rubisco) also accepts O2 in its active site which can decrease PSS by up to 50%. This is because the enzymeevolved in a non-O2 atmosphere. This wasteful process is called photorespiration and has nothing to do with mitochondrial respiration.It is an extremely limiting factor in the productivity of plants such assoybeans, wheat, oats, barley under optimal conditions.

-C4 plants such as corn, sorghum, sugarcane overcome photorespiration by a strategy that concentrates CO2 at the site of photosynthesis resulting in up to 3X higher rates of PSS compared to wheat, rye,barley, oats, soybeans and all other dicots.

-Enhanced CO2 in maize also allows plant to reduce stomata opening, thus conserving H20 and allowing these plants to grow in hot climates-by contrast,another tropical plant, rice, did not evolve a CO2

concentrating strategy and can only grow in hot climates when thereis lots of water. Slide 10.11

8. Using 3C molecules How are large-structures derived from these 3C molecules?

The 3C can be broken down to a reactive form of 2C subunits (acetyl-CoA) which can be joined together to form fats/oils.

The 3C can be joined to 3C to form 6C, such as glucose. Chains of glucose can form cellulose (cell wall) or starch.

6C can be broken down to 5C, ribose, which is a major part of DNA/RNA.

Different C-C combinations can join together to help form the backbone of amino acids (along with N, S, etc.) and all the other molecules of the cell.

How can energy be derived from the 3C molecules?3C is broken down to 2C, which enters the mitochondrion. There, the C-C bonds are broken to create 2 x CO2 which is then released. This releases energy that is again used to generate cofactors (ATP, NADH).

This process is called respiration and it requires O2 to accept electrons (which had released energy as they were transferred through a series of reduction-oxidation reactions). In the presence of 4H+ protons, the reduced O2 generates 2H20.Therefore, what are the different fates of the 3C molecules derived from the Calvin cycle?

Slide 10.12

Overview of Carbon + Energy Assimilation & Flow

H20

Electrondonor

02ElectronExcited

SolarEnergy

High energy electronmolecules (NADPH, ATP)

Electron/energy used to create Carbon-carbon additions (glucose, C6)

Carbon-carbon bonds broken to create energeticbuilding blocks (eg. 2C, acetyl-CoA) to build newcarbon-based organic structures and ATP topermit exchanges of electrons for biochemistry

CO2

•3C sugars allow transfer between cellular compartments•Sucrose (6C + 6C) is the form for source-sink transport (sap)•Starch (6C+6C+6C….) is the long-term storage form of Carbon

•a 3C sugar is the initial product

Slide 10.13

Leaf Cell =Photosynthate Source

Xylem Phloem

Cytoplasm

Photosynthate sink cell (seed, pollen, roots, fruits, growing tissues)

H20

H20

Roots

Sucrose

Sucrose

Sucrose

Stomate

Atmosphere

CO2O2

Light Energy

CHLOROPLAST

Chlorophyll Complex

H20 O2

High energyelectrons

ADP ATPNADP NADPH

MITOCHONDRION(Respiration)

Calvin Cycleuses energyto build C-Cbonds

CO2RubiscoEnzyme

Leaf air space

5Cacceptor

2x 3C

3 turns =3C

5C

ATP ADP

NADPH

NADP

3C 3C

3C + 3C=6C Glucose

3C Pyruvate

Glycolysis (6C to 3C)regeneratesNADH + ATP

3C Pyruvate

6C Glucose + 6C Fructose =12C Sucrose

Oil bodies(oil, fatty acid store)

Mitochondrion (releases energyfor growing tissues)

Amyloplasts(starch store)

Citric AcidCycle (Krebs)releases energyby breakingC-C bonds

ATPNADH

2C Acetyl-CoA

4C acceptor

6C citrate

4C

CO2

NADH(lots)

ATP(lots)

O2H20

Transpiration(H20 evaporation)

3C + 3C=6C Glucose

6C + 6C + 6C....=Starch (storage)

2C Acetyl-CoA

6C glucose + 6C

3C Pyruvate

Glycolysis

Citric AcidCycle (Krebs)releases energyby breakingC-C bonds

CO2

2C

6C + 6C + 6C....=Starch (storage)

3C

2C Acetyl-CoA

2C Acetyl-CoA

Building blocks forcomplex carbonmolecules (fattyacids, amino acids, etc.)

Building blocks forcomplex Carbon molecules

Building blocks

Break-downRecycling of 2C

PhotorespirationRubisco acceptsO2 = 50% carbon

waste. C4 plantsconcentrate CO2

H20

2C + 2C + 2C.....=18C, 20C fats, oils

source: M. Raizada