Botany 2104 - Plant Form and Function

Respiration Overview

Cellular (Aerobic) Respiration
The overall reaction for respiration:

C6H12O6  +  6 O  + 6 H2O   ------------------------>  6 CO2  +  12 H2O  

Basically, the process involves oxidizing sugar carbons to CO2.  An electron transport system is used to create a H+ (proton) gradient that is used to make ATP.  The final depository for the electrons is O2, which is reduced to water.

Aerobic respiration is strongly identified with the mitochondria, however, the starting reactions are in the cytosol.  If oxygen is available, the mitochondria get involved with respiration, and the internal structure of the mitochondria is critical to the process.

Mitochondria
Two membranes:  outer membrane and inner membrane
Small volume aqueous compartment:  intermembrane space
Large volume aqueous compartment:  matrix

The carbon oxidation takes place in two sets of reactions, glycolysis in the cytosol and the Krebs cycle in the matrix of the mitochondria.  Both glycolysis and the Krebs cycle occur in steps.  Stepwise oxidation is important because:
1.  it allows the dissipation of energy that is released as heat
2.  it generates intermediates ==>  steps to start making amino acids, N-bases, other sugars for cell wall and nucleic acids, fatty acids, chlorophyll, anthocyanins, hormones, secondary metabolites, etc.

Glycolysis
Glycolysis oxidizes glucose or fructose (C6 sugars) or triose-P (C3 sugars) to pyruvate (C3 acid).  Some of the energy released during the oxidation can be captured as ATP.  The electrons released by the oxidation are held by NADH.  Pyruvate is an organic acid.  If the carbon is organic, it is at least partly reduced.  Therefore, there are still calories (energy) available in pyruvate.  

The Krebs cycle
The Krebs cycle finishes the carbon oxidation process.  All of the carbons in pyruvate are oxidized to CO2.  It is a cyclic process, with the pyruvate carbons getting attached to an existing cycle molecule prior to oxidation.  Once the pyruvate carbons are completely oxidized, the cycle molecule needs to be regenerated.  A little bit of ATP is made during the Krebs cycle, but mostly it is an oxidation process.  Most of the electrons are held by NADH.  A few are held by FADH2.

Electron Transport Chain (ETC)
The electrons from NADH and FADH2 are passed to a system of electron carriers in the inner mitochondrial membrane, eventually reaching O2 which is then reduced to H2O.   As the electrons pass through the carriers, protons (H+) are moved from the matrix to the intermembrane space.  This establishes a proton gradient.  When the protons pass through a channel protein back to the matrix, ATP is synthesized.  How much ATP gets made?  It depends on who is doing the counting.  Most plants that oxidize hexose molecule completely to CO2 will get between 30 and 38 ATP from glycolysis, Krebs cycle, and ETC.  The majority of that ATP (85-90%) comes from the ETC.   The high end of ATP yield (35-38 ATP) represents about 40% of the energy that was available in a molecule of glucose.  The remaining energy is given off as heat.

Some plants deliberately have an ATP yield in the mid-teens.  Their electrons take an alternative route in the ETC and don’t build as large as proton gradient by the time they get to O2.  Because the ATP yield is down, these plants release more heat.  These plants are called thermogenic.  They can raise the temperature in their microenvironment as much as 10oC.  This temperature increase volatilizes scents to attract pollinators more efficiently.

Anaerobic respiration
Plants are able to respire anaerobically – without oxygen – for a short time.  Glycolysis still occurs, but any reactions involving the mitochondria are not available.  Without oxygen and the ETC, where can NADH unload its electrons?   Enter fermentation:  a set of reactions that let NADH unload electrons so glycolysis can continue.   Most plants use alcohol (ethanol) fermentation, but some use lactic acid fermentation, and some use both types. The most likely anaerobic scenario for plants  ==>  roots in water-logged soil.

Some plants (hydrophytes) are specialized to have their roots always in water, like anchored aquatic plants and emerged plants (water lilies, cattails, etc.)  The internal anatomy of these plants has large air spaces that form passage ways to get O2 to the roots.


Respiration - Details

Respiration is essentially the cellular burning of a fuel to release its energy.  To burn the organic matter in a log, we would strike a match to ignite the log and release the available energy as heat.  In respiration, the starting carbohydrate must be ignited; some of the released energy will be as heat, but the rest of the energy will be captured as ATP.  The cellular equivalent of the match is ATP.  The cell will have to spend some existing ATP to start respiration, just as we need to use an existing flame to ignite the log.
 
Glycolysis
The standard depiction of glycolysis is given in Figure 1.  The process starts with glucose.  Two ATP are used to “ignite” each glucose molecule.  The end result is two molecules of triose-P, dihydroxyacetone phosphate (DHAP) and phosphoglyceraldehyde (PGald).  Energy has been expended, but no energy has been collected.  All of the carbon is present as carbohydrate, which was the starting form of the carbon.
The next steps of glycolysis will oxidize the sugar carbons.  The oxidation process will release energy.  Some of that energy will be captured as 4 ATP molecules.  Some of the energy will be lost as heat.  The remainder of the released energy will still be potentially available.  That last portion of energy is associated with the electrons being held by 2 NADH molecules.

In general, plants don’t have a lot of free glucose lying around their cells.  Most of the sugars to be respired are in sucrose (arriving via the phloem), newly made triose-P exported from the chloroplasts,  and stored carbohydrate polymers such as starch and fructans.  Figure 2 shows how plants might get carbohydrate from these sources to the triose-P step of glycolysis.  Plants don’t necessarily need to spend ATP to start the oxidation of sugars.  Glucose can be phosphorylated as it is released from starch by the action of starch phosphorylase.  There is an alternate enzyme for converting fructose-6-P to fructose-1,6-bisP that uses inorganic pyrophosphate instead of ATP.

Also, plants don’t necessarily make ATP during the oxidation steps of glycolysis.  It all depends on their phosphate status.

The reality of ATP yield for plants from glycolysis of one molecule of glucose equivalent:  -2 to + 4.

If oxygen is not present, the pyruvate that remains at the end of glycolysis will be fermented.  Fermentation allows the cell to unload the electrons that were given to NAD+ during the oxidation steps of glycolysis.  This will free up the NAD+ molecules to get more electrons and keeps glycolysis going.  As fermentation is essentially a carbon reduction step, no additional energy is released or captured as ATP in the process.  The possibilities for fermentation in plant cells are shown in Figure 4.

If oxygen is present, the oxidation of the carbons in pyruvate to CO2 continues in the matrix of the mitochondrion.  This is where the Krebs cycle (TCA cycle, citric acid cycle; Figure 5) is located.  Pyruvate is initially oxidized to release one carbon as CO2 with a C2 acetyl group to be attached to coenzyme A (CoA).  The oxidation step also results in the reduction of NAD+  to NADH.  The acetyl group is joined with an existing molecule of oxaloacetate (C4) to give citrate (C6).  Citrate is oxidized to α-ketoglutarate (C5), with the reduction of NAD+  to NADH and the release of one carbon as CO2.   α-Ketoglutarate is converted to succinate (C4), with the release of one carbon as CO2 and the phosphorylation of one ADP to ATP.  Succinate is oxidized to fumarate (C4), with the reduction of FAD to FADH2.  Following the conversion of fumarate to malate (C4), the malate is oxidized to oxaloacetate (C4), with the reduction of NAD+  to NADH  The cycle is now reset to take in an acetyl group from the next pyruvate.

Summary of the Krebs cycle:
pyruvate + ADP + Pi + 4 NAD+ + FAD  -------->  3 CO2 + ATP + 4 NADH + FADH2
The Krebs cycle happens twice per glucose equivalent that started glycolysis.

Krebs cycle + glycolysis:
1.  hexose ------>   6 CO2
2.  phosphorylation of 0-6 ADP to 0-6 ATP
3.  The oxidation of hexose released 24 electrons, now being held by 2 NADHG, 8 NADHK, and 2 FADH2.  (The G and K subscripts on NADH indicate that these molecules were reduced during glycolysis and the Krebs cycle, respectively.)

Notice that aerobic respiration, which required molecular oxygen, hasn’t used any oxygen yet.  The ETC (Figure 6) will transport the electrons from the dinucleotides to O2.  The O2 will be reduced to water.  
    6 O2  +  24 electrons  +  24 H+  -------->  12 H2O

As the electrons move through the ETC from the dinucleotides to O2, a proton gradient is established.  A high concentration of protons builds up in the intermembrane space.  These protons will diffuse through an ATPase located in the inner membrane of the mitochondrion.  As the protons pass through the ATPase, its configuration changes.  The enzyme is then able to attach a free phosphate to an ADP, forming ATP.

The steeper the proton gradient, the more ATP that can be made.   Figure 6 shows that protons can pass from the mitochondrial matrix to the intermembrane space at three points in the ETC.  Only electrons that are obtained from NADHK and given to Complex I will travel through all three proton passage points.  Electrons from NADHG and FADH2 only travel through two proton passage points.   Various calculations and measurements show that a maximum of three ATP are made per NADHK  and two are made per NADHG  or FADH2.   The actual number of ATP made depends on many factors, such as cellular demands for reducing power elsewhere, whether NADHK  gives its electrons to Complex I or NADH dehydrogenase (B in Figure 6),  and whether or not the alternative oxidase is used.

Inhibitors of the ETC:
Complex I:  rotenone, piericidin
Complex II:  TTFA (thenoyltrifluoroacetone)
    (succinate —> fumarate:  malonate)
Complex III:  antimycin
Complex IV:  CO, CN-, N3-

Uncouplers:  collapse the proton gradient.  Oxygen is still reduced to water, but no ATP can be made.  dinitrophenol (DNP)

Figure 7 is an abbreviated version of glycolysis and the Krebs cycle.  These two pathways generate many intermediate molecules as hexose is oxidized to CO2.  At any point in the pathways, the intermediates can be removed to make other molecules that the plant needs, including both primary and secondary metabolites.


Review

Respiration
    aerobic vs anaerobic
    location of processes:  cytosol vs mitochondria
    purposes of respiration
    why stepwise oxidation is important
    glycolysis:  What does it accomplish?
    fermentation:  Why is this process important to plant cells?
    Krebs cycle:  What does it accomplish?  What is its importance beyond pyruvate oxidation?
    electron transport chain:  What does it accomplish?  How do electrons reach the chain?  How do plant mitochondria get around blockages?

Glycolysis and the Krebs cycle are important not just as a means of carbohydrate oxidation, but also as a source of intermediates.  What is meant by the term intermediate in this context?  Name three types of molecules that can be constructed from the intermediates of glycolysis and  the Krebs cycle.


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27 July 2011