READINGS:

 

a)      REQUIRED:

 

1-     Slattery, C.J., Kavakli, I.H., Okita, T. W. 2000. Engineering starch for increased quality and quantity. TIPS 5#7:291-298.

2-     Trethewey, R.N., Riesmeier. J. W., Willmitzer, L., Stitt, M., Geigenberger, P. 1999. Tuber-specific expression of a yeast invertase and a bacterial glucokinase in potato leads to an activation of sucrose phosphate synthase and the creation of a sucrose futile cycle. Planta 208: 227-238.

 

b)      OPTIONAL:

 

1-     Douce, R., Neuberger, M. 1989. The uniqueness of plant mitochondria. Ann. Rev. plant physiol. Plant Mol. Biol. 40:371-414.

 

 

 

 

RESPIRATION IN PLANTS

 

 

-         DEFINITION: Respiration is the process whereby reduced organic compounds are mobilized and oxidized in a controlled manner to yield useful forms of energy.

 

 

-         Complex organic molecules such as glucose contain much potential energy because of their high degree of structural order (so little entropy).

 

 

 

-         Free energy is high when entropy is low, and this can be found from the combined expression of the first and second laws of thermodynamics:

 

 

 

 

 

-         Various biological compounds have significant DG^o` but ATP, PEP, etc, have relatively high DG upon only transfer of the phosphoryl group. See tables 14-2 and 14-4.

 

 

 

 

 

 

-         The ATP- ADP cycle:

 

 

 

ATP is a carrier of energy-rich phosphoryl groups from reaction of catabolism to reactions of anabolism. In plants this energy is used for transport and biosynthetic work, but also mechanical work, i.e. MT motors. In animals, muscle work is a major consumer.

 

 

 

 

 

 

 

 

-         Nicotinamide adenine dinucleotide cycle:

 

NAD, NADP carry energy-rich electrons from catabolic reactions to electron-requiring anabolic reactions. NADPH+H primarily for biosynthetic reactions, NADH+H primarily for electron transport.

 

 

 

-         The overall gas exchange and free energy change for the complete oxidation of a mole of glucose is:

 

 

C₆ H₁₂ O₆ + 6O₂ + 6H₂O à 6CO₂ + 12H₂O ΔG -686 kcal

 

 

-         The respiratory quotient (RQ) is:

-          

The molar ratio of CO₂ consumed.

 

The RQ and the amount of ΔG depend on the type of “fuel” respired.

 

Respiratory quotient and energy yield of major biological fuels
Energy product, kcal
RQ	Per gram of fuel	Per liter of O2
More highly oxidized than carbohydrates: Organic Acids 1.30   Carbohydrates 1.00	--   4.18	--   5.05
More highly reduced than carbohydrates: Triacylglycerols 0.71   Proteins 0.80	9.46   4.32	4.69   4.46

 

 

 

-         Respiration serves two major functions:

 

 

-         1) To provide ATP and NADH needed for maintenance reactions and growth.

 

-         2) Provide carbon skeleton for the synthesis of metabolic intermediates, both primary (amino acids, protein, nucleic acid, TCA, storage biosynthesis, etc.) and secondary (terpenes, phenylpropanoids, isoprenoids, flavanoids).

 

-         Not all of the carbon that enters respiration is converted completely to CO₂ and H₂O. While glucose is commonly cited as main respiratory fuel, sucrose, starch, fructosans, other sugars, lipids, organic acids, etc. can be used as illustrated in fig. 13-12.

 

 

 

-         Respiration is a multi-step process:

 

Stage	Location	ATP Synthesis Mode
1) Glycolysis (most primitive form of carbon catabolism	Multi-enzyme complex on cytosolic face of outer mitochondrial membrane. Segments may move about. Also, in chloroplast.	Substrate level (does not involve a proton gradient)
2) TCA Cycle (in mito)	Matrix (lumen) of mitochondria. Probably as multi-enzyme complex.	Substrate level
3) Electron transport to 02 (in mito)	Inter membrane of mitochondria	Powers ATP synthesis
4) Utilization of the PMF produced as a result of electron transport (in mito)	CFo/CF1, of inner membrane	Major source of ATP

 

 

 

-        

Glycolysis:

 

 

 

The overall reactions of glycolysis take place in two major stages:

 

1)      In the first stage 1, 6-carbon molecules are raised to a higher energy level at the expense of ATP (priming).

 

2)      In the second stage the products of stage I are converted into 3-carbon, pyruvate molecules with conservation of energy as ATP and NADH. The net yield of conserved energy is 2 ATP and 2 NADH per glucose.

 

 

-         The glycolytic sequence is known to occur in both the cytosol (on outer mito membrane) and the chloroplaste stroma in plants.

 

-         We do not have time to consider energy yields of glycolysis. You may remember some of this from earlier Biochem courses.

 

 

-         Glyconeogenesis (reverse glycolysis- sort of):

 

The operation of glycolysis in reverse - sort of! This metabolism is particularly prominent during germination of fat (oil) rich seeds (e.g. castor bean, sunflower). You will remember from the cell biology lectures that the glyoxylate cycle in the glyoxysomes takes fatty acids hydrolyzed from TAG’s in the lipid bodies and converts them to succinate. The mitochondria convert succinate to malate. In the cytosol the malate is converted to oxaloacetate and this to PEP. PEP is then converted to sucrose by gluconeogenesis. The sucrose is then used to support growth of the seedling until PS can take over:

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

-         But, remember that the irreversibility of phosphofructo kinase stands in the way of PEP à sucrose. So, both plants and animals circumvent this by the fructose-bis-phosphatase enzyme:

 

 

 

 

 

 

 

-         This costs the energy that was added during “ priming” of fructose-6-PO₄ but it allows reversal. However, note that F-2, 6-bis PO₄ strongly inhibits FBPase. (Ann.Rev. Plant Physiol. Plant Mol. Biol. 41:153-185, 1990; Ann. Rev. Bch. 64:799-835, 1995).

-          

-         F-2, 6-BP is a signal metabolite that is:

1)      Found in all eucaryotes at 1-10µM.

2)      An activator of PFK in animals (but not in plants), so is a

glycolytic signal that can increase FBP.

3)      An inhibitor of FBPase in plants.

4)      Tied to sucrose synthesis in that (in plants) increased cytosolic F-2,6-BP decreases sucrose synthesis, which indicates that gluconeogenesis is strongly regulated by F-2,6-BP.

 

-         Complex regulation of glycolysis:

 

 

 

 

 

 

 

-         Gluconeogenesis- way around the:

 

1)      Hexokinase block:

 

G-6-PO₄ + H₂O à Glucose + Pi

 

Also, F-6-P à G-6-P à G-1-P à UDP-G à Sucrose-P à Sucrose

 

G-1-P à ADP-G à Starch

 

So the Hexokinase block very important in plants

 

2)      PFK block:

 

F-1,6 BP à F-6-P + Pi

Or reversible of PFP reaction

 

3)      Pyruvate kinase block:

 

Complex series of reactions (mito.) animals.

 

In plants PEP carboxykinase allows

 

OAA + ATP à PEP + ADP + CO₂

 

 

 

The synthesis of F-2,6-BP is tied to triose-PO₄(Pi) levels as shown below. How cytosolic triose-PO₄/ PO₄ ratio promotes synthesis of F-2,6-BP, which in turn inhibits the hydrolysis of F-1,6-BP to F-6 and Pi. The resulting lower F-6 P concentration reduces the rate of sucrose synthesis.

 

So all 3 blocks to glycolysis easier in plants than animals. Why?

 

 

 

 

 

When photosynthesis increases in response to increased light or CO₂, 3-PGA and triose-P levels increase and Pi levels presumably decrease. These changes would inhibit formation of F-2,6-BP and stimulate its turnover (3 to 5- fold decrease). Inhibition of ATP or NADPH synthesis results in a decrease in 3-PGA and other triose-P and an increase in F-2,6-BP.

 

-         Glycolysis, gluconeogenesis, sucrose synthesis, and starch synthesis are highly regulated in plants. Shown below are some key enzymes and their regulation (all irreversible reactions):

 

1)      Hexokinase: Glucose + ATP à G-6-PO₄ + ADP

 

(-) by G-6-PO₄.

2)      Phosphfructokinase (PFK): F-6-P + ATP à F-1,6 BP + ADP

 

(-) by high conc. of ATP, NADH, citrate, PEP.

(+) by ADP.

 

3) Pyruvate kinase: PEP + ADP à Pyr + ATP

 

 

(-) by high conc. of ATP

(+)F-1,6 BP and PEP.

 

4)      Fructose 1,6 bisphosphatase: F-1,6 BP à F-6-P + Pi

 

(-) AMP

(+)F-2,6 BP

 

5)      Fructose-6 BP kinase: F-6-P + ATP à F-2,6 BP + ADP

 

(-) PPi

(-) 3-PGA

(+) Pi

(+) F-6-P

 

6)      Fructose 2,6 bisphosphatase: F-2,6 BP à F-6-P + Pi

 

(-) Pi

(+) 3-PGA

 

7)      Sucrose phosphate synthase: F + UDP glucose à Suc-6-P + UDP

 

(-) Pi

(+) G-6-P

 

-         Pi regulation is extensive in plants because Pi is usually deficient in the environment and Pi is critical for many processes.

 

 

A model depicting alternative pathways of glycolytic carbon flow and mitochondrial respiration:

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

Plants (but not animals) have yet another enzyme to reverse the PFK block. This is PPi- dependent Phosphfructokinase (PFP) or PPi linked PFK. It catalyzes the reversible reaction:

F-6-P + PPi ßà F-1,6 BP + Pi