Bioenergetics and Metabolism

Part II and Chapter 13

Bioenergetics and Reactions

– Thermodynamics applies to biochemistry – Organic chemistry principles at work – Some biomolecules are “high energy” with

respect to their hydrolysis and group transfers

– Energy stored in reduced organic compounds can be used to reduce cofactors such as NAD+ and FAD, which serve as universal electron carriers

Key topics: Learning Goals

Metabolic Pathways Cooperate To:

1. Obtain Chemical Energy by:a. Capturing Solar Energy, or b. Oxidizing Energy Rich Chemicals from the Environment.

2. Convert Nutrient Molecules to metabolic intermediates, then monomers or waste products.

3. Polymerize monomers to polymers (proteins, carbohydrates, nucleic acids, lipids).

4. Synthesize and Degrade (turnover) biomolelcules.

Anabolism and Catabolism

Linear and Circular Pathways

Metabolic Pathways

Auto-Pathways

Pathways Arranged as Multi-Protein Modules

Flagella

LPS

Outer Membrane

Peptidoglycan

Cytoplasmic Membrane

Glycolysis

ATPase

RNA

5 Main Classes of Metabolic Reactions

1. Oxidation-Reduction Reactions

2. Reactions that Make or Break Carbon-Carbon Bonds

3. Internal Rearrangements, Isomerizations, Eliminations.

4. Group Transfer Reactions.

5. Free Radical Reactions.

Chapter 13 Bionergetics ATP

Showed that Respiration Was Oxidation of Carbon and Hydrogen…thus began Thermodynamics

Laws of Thermodynamics

First Law – for any change, the energy of the universe remains constant; energy may change form or it may be transported, but can not be created or destroyed.

Second Law – The Entropy Law can be stated 3 ways:

1. Systems tend from ordered to disordered.

2. Entropy can remain the same for reversible processes but increases from irreversible processes.

3. All processes tend towards equilibrium.

Everything Equilibrium = Death.

Third Law – Entropy goes to zero when ordered substances approach absolute zero = 0oK

Thermodynamics

Gibbs Free Energy G and ΔG

Enthalpy H and ΔH

Entropy S and ΔS

ΔG = ΔH - TΔS

Biochemistry Uses ΔGo’ Not ΔGo

Standard Conditions (all reactants and products at 1M, gases at 1 atm, Temp = 25C) are Not Biological

Conditions

So, ΔGo’ takes out water (55.5M), and [H+] is set at pH 7 (not 1M which would be pH=0)

and for humans ΔGo’ uses 37oC (310 K), but for bacteria ΔGo’ uses 25oC (298 K)….or the temperature

of the environment.

ΔGo’ = - RT ln Keq

You should be able to do EOC Problems 2 and 3 easily

EOC Problem 6: the difference between ΔGo’ and ΔG.

Free energy, or the equilibrium constant, measure the direction of processes

ΔGo’s Are Additive

Hexokinase Rxn: Glucose + ATP Glucose-6-P + ADP

Glucose + Pi Glucose-P + H2O ΔGo’ = 13.8 kJ/mole

ATP + H2O ADP + Pi ΔGo’ = -30.5 kJ/mole

Overall = ΔGo’ = -16.7 kJ/mole

Exergonic ! So:

K’eq = 7.8 x 102

EOC Problems 9 and 12: the ΔGo’ for 2 coupled reactions.

Biochemical Pathways Have Evolved To:

1. Use reactions that are relevant to metabolic systems:

Makes use of available substrates – with reaction rates that are NOT slow (have too high activation energies even with enzymes!) to produce useful products (which are themselves substrates). And,

2. Maximize Rates

Evolution’s Toolbox: reactions that work. : circumvent “impossible”

reactions. : most reactions in organic

chemistry occur in biology, except one, the Diels Alder Rxn…but we will see about that.

You be a radical !

You be inonic !

Rich in electrons donate electrons

Electron poor suck up electrons from donors

The Importance of Carbonyls

Nucleophile

Electrophile

Imines are like carbonyls

Here the carbonyl is an electrophile

Making and Breaking Single Bonds

Isomerations are Internally Complex

The Classic Redox Reaction

ATP Hydrolysis

Energy Charge

Energy Charge =

[ATP] + ½ [ADP] [ATP] + [ADP] + [AMP]

Energy Charge

Why the ½ [ADP] ??? It is because of

Adenyl Kinase:

ADP + ADP ATP + AMP

Nucleotide Intracellular Concentrations*

Nucleotide Conc, μM Nucleotide Conc, μM ATP 3,000 GTP 923

ADP 250 GDP 128

AMP 105 GMP 20

dATP 175 dGTP 122

dTTP 77 dCTP 65

UTP 894 CTP 515

cAMP 6 cGMP nd

ppGpp 31

NAD+ 790 NADP 54

NADH 16 NADPH 146

FAD 51 FMN 88

AcCoA 231 SuccCoA 15

in Salmonella enterica subsp Typhimurium

from Bochner and Ames, 1982, J. Biol. Chem 257:9759-9769

Magnesium Stabilizes Tri- and Di-phosphates

EOC Problem 19: How much ATP is used in a human/day.EOC Problem 20: About turn over of the α and β phosphates (can you located them above?).

Pyruvate Kinase

1,3-Bisphosphoglycerate has More Energy Than ATP

Phosphocreatine Is Store of Energy in Muscle

What About Actual ΔG ?

ΔG = ΔG’o + RT ln([products]/[substrates])

This is the real, biological ΔG in a cell !!

At 25oC RT = 2.48 kJ/mole (2.5 kJ/mole)

At 37oC RT = 2.58 kJ/mole (2.6 kJ/mole)

We will be doing this a lot later on !

Doing Worked Example 13-2

Using E. coli

ΔG = ΔGo’ + RT ln [ADP][Pi]/[ATP]

ΔG = -30.5 kJ/mole + [ (8.315 J/mole.K)(310K) ln(1.04mM)(7.9mM)/7.9 mM]

ΔG = -30.5 kJ/mole + 2.58 kJ/mole (-6.8)

ΔG = -30.5 kJ/mole + (-17.6)

ΔG = -48.1 kJ/mole

Note: Calculate mM such as 1.04mM = 1.04 x 10-3M

In the text for the Human Erythrocyte it works out to ΔG = -52 kJ/mole

Acetyl-CoA (Thiol-ester) Has the Energy of ATP!

EOC Problem 21: Cleavage of ATP to AMP + PPi…..why is this different (see Table 13-6 above). (What DNA enzyme did the same? It’s in Chapter 8)

Enzyme Reaction Phosphorylation Intermediates Used to form C-N Bonds

Phosphates: Ranking by the Standard Free Energy of Hydrolysis

Reactions such as

PEP + ADP => Pyruvate + ATP

are favorable, and can be used to synthesize ATP.

Phosphate can be transferred from compounds with higher ΔG’ to those with lower ΔG’.

Nucleoside Diphosphate Kinase makes NTP’s from ATP and NDP’s

Carbon Redox – Watch the Red Dots (Electrons)

Emf or Eh or Eo

EOC Problem 24: Respiratory chain thermodynamics (we will do this in Chapter 19)…learn it well now!

Calculations

Differences between half cells…Example of electron transfer from NADH to cytochrome-b:

NADH Eo’ = -.32 v

Cyt-b Eo’ = 0.077 v

ΔEo’ = Eo’oxidized – Eo’ reduced = 0.077v – (-0.32v)

ΔEo’ = 0.397v

Further Calculations

What is the ΔG’o for oxidation of NADH by cytochrome-b

ΔG’o = - nℱ ΔEo’ Faraday Constant = 96,480 J/v.mole

ℱ = 96.5 kJ/v.mole

ΔG’o = - (2) 96.5 kJ/v.mole (0.397v) = - 77 kJ/mole

ΔE = ΔE’o + (RT/nℱ) ln ([products]/[substrates])

EOC Problem 25 and 26 are all about this.

What about the real ΔE ?...and then ΔG !

NAD+ + 2e- + 2H+ NADH + H+

Lactic Acid Dehydrogenase = LDH

Rossmann fold, a structural motif in Dehydrogenases

Vitamin Niacin is Made from W and Needs to be Amidated for NAD+

FMN and FAD

Enzyme Reactions have a Yield of ~1.0The Perfect Catalysts

Assume Metabolism worked on each step in the metabolic pathway having a yield of 0.9 (tremendously high for organic chemistry reactions!!).

Then look at a 10 reaction pathway such as Glycolysis:

if you start with 100 mg of glucose the pathway would only produce less than 39 mg of pyruvate….AND, the cell would fill up with 61 mg of side reaction products!

The message is that metabolism and life would be tremendously inefficient = motionless, wasteful BLOBS filled with junk.

Most enzymes have a yield of 0.9990 to 0.99990 (that is they make a mistake reaction 1 in a 1,000 to 10,000 reactions).

What is the BEST Yielding Enzyme?

Answer: DNA polymerase makes an error 1 in 107 to 109 reactions.

Why? It is the one of the few enzymes to have a “proof- reading” function to correct the 1 in 103 to 104 mistakes.

Check it out in Molecular Biology!

Things to Know and Do Before Class*

1. The basic laws of thermodynamics.

2. Be able to calculate ΔG, ΔGo’ from concentrations or Keq.

3. Be able to calculate over all ΔGo’ from summed reactions.

4. Principles that make some bonds “high energy”.

5. EOC Problems (2, 3), 6, 7, 9, 12, 14, 20, 21, 24-26.

*Two Class periods for this chapter.