Introduction to Metabolism

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Biochemical Energetics and MetabolismChem114 Spring 2015Prof. Roy WollmanGo over syllabusGoals and overviewCommunication RulesSchedule (order of classes NOT the same as the book)Interactive learning. Thinking exercises in class:

In-class multiple choice

No Points.

Help you and me to see that we all understand the materialOpen questions and group discussions.

These will be questions that will require some thinking. In some cases I will give 2-3 min for group discussionsWhich surfboard is good for beginners?

FishFunboardLongboardAll of the above. Surfing is really easyKey three question you probably have: Why study Metabolism?

What is Metabolism?

How to study Metabolism?Why study Metabolism?

What cause obesity?

Eating habits?

High fructose corn syrup?

Too much TV?You are what you eat

?

Metabolic engineering

Why study metabolism?Major health challengeTry to do something about it.Understand where it is coming from

Major technology for (almost) all the non-biomedical biotechnology companies. Designer organismSustainable, green, efficient chemical productionWhat is metabolism? What is a car? What is a car?

Is this a good representation of a car?

Is this a good representation of a car? What information do you need to know to understand how a car works?

We will talk about it as a class in 2-3 minutes

Turn around to your neighbors and discuss the following question: What information do you need to know to understand how a car works?List of parts (engine, cylinder, etc.)

How each part work.

How all the parts work together. What is metabolism?

List of compounds and reactions (metabolites, enzymes, co-enzymes, )

Mechanism of action for all of these reactions

Regulation that makes it all work in unisonHow to study Metabolism?Metabolic Vocabulary: Learn about the enzymes and metabolites that build up key metabolic networks. Metabolic Grammar: Learn about the chemistry that of metabolic reactions that create metabolites. Metabolic Poetry: Learn about the integration of metabolic reactions into metabolic networks. How to study Metabolism? Q: Do I need to memorize stuff? But GoogleA: I cant explain to you how a car works if you need to google what an piston is every second.Q: Do I need to think about stuff? But I can memorizeA: YES! To understand how it all connects you will need to challenge you understanding. Learning is YOUR responsibilityLearning is YOUR responsibilityRead the assigned reading to help with the terms, vocabulary etc. Make sure you are not getting lost. Thinking exercises in class

How to study Metabolism?

Key three question you probably have: Why study Metabolism?

What is Metabolism?

How to study Metabolism?

Chemical principles of metabolism: Enzyme catalysis and co-factors

Thermodynamics and metabolic flux.

Catabolism & Anabolism

Enzymes provide unique chemical environment

Vitamin deficiency disorders

Co-factors help enzymes to create the required unique chemical environment

Thermodynamics of chemical reactions

Enzyme catalysis makes things move faster

Gibbs free energy:

A + B C + D

Thermodynamic potential: Energetic potential of this SPECIFIC reaction given reactants and products concentrationEnergetic potential

= 6 kJS + Q P + RSSSSSSSSQQQQQPPRR[S][Q][P][R]= 10

= 6000 - ln(10) * 8.3145 * 310 0

Can eating broccoli change ?

YesNo Maybe Longboard

Metabolic pathway are irreversible

Game: You are a metabolic engineer

You want to break down glucose and make ATP.

You have -100 kJ/mol of free energy to distribute over the four reaction.

How will you do so?

Turn around to your neighbors and discuss the following question:

-90, -4, -3, -3 -3, -4, -3, -90 -25, -25, -25, -25 -5, -40, -40, -5

How will you distribute -100 kJ/mol over the four reactions? Commitment!An irreversible (exergonic) reaction will commit the cell to generating the product.

This helps drive the entire pathway even if many of the steps are close to equilibrium



Why use different routes?

Chemical principles of metabolism: Enzyme catalysis and co-factors

Thermodynamics and metabolic flux.


Principles of metabolic flux regulationAllosteric regulation and feedbacks

Covalent modification

Genetic control (expression & isozymes)

Substrate cycles

Allosteric regulation and feedbacks

What can you do with negative allosteric regulation?


Allosteric regulation and feedbacks

Which reaction is the most likely to be negative allosteric regulated?

Regulation through covalent modification

Phosphorylation allows for external regulation of the metabolic networks

Gene expression

Why have three different regulatory approaches:

Turn around to your neighbors and discuss the following question: Allosteric regulation and feedbacksCovalent modificationGenetic control (expression & isozymes)

Hint: Which one is the fastest? which one is the slowest? Or are they all happening at the same time?Substrate cycle

If we increase f and reduce r the overall flux changes dramatically.

If it was a single enzyme doing both reaction, increasing its activity would cancel out. e


18 fold increase 1.8 fold increase 9 fold increase LongboardIf we can increase the rate of f by 80% and reduce the rate of r by 90% through addition of a phosphate group. What is the gain we will see: 1.80.1= 18Principles of metabolic flux regulationAllosteric regulation and feedbacks

Covalent modification

Genetic control (expression & isozymes)

Substrate cycles

The Currencies of energyATP

Other Phosphorylated compounds

Coupled reactions

Theioesters

ATP, ADP, AMP, & Adenosine54What can you do with positive allosteric regulation?


What will I get from the hydrolysis of 1 gram of ATP at room temperature?

[ATP] = 3 mM, [ADP] = 0.8 mM, [Pi] = 4 mMR=8.3145 J/K, T=310 K, ATP molecular weight is 507.18 g/mol1 gram ATP 2 mmol

So 1 gram ATP will give us 96.2 J2 C8H18+ 25 O2 16 CO2+ 18 H2O42,400J/ g

50 J96.2 J 96.2 kJ LongboardATP is the energy currency of the cell

$$

Phosphoryl groups

How to spend biochemical money?

Hexoinase: an enzyme that catalyzes the addition of a phosphate group to glucose

PiH2O

+13.8 kJ/mol-13.8 kJ/mol ~0 kJ/mol LongboardWhat is the DG of this reaction? How can a cell create G6P?

Use an enzyme that will catalyze the reaction faster despite the positive DGLongboard Use special co-factors that will change the chemical environment and will make this reaction exothermic Couple this reaction to another exothermic reactionCoupled reactions:

Hexoinase: an enzyme that catalyzes the addition of a phosphate group to glucose

What happens if a reaction endothermic with DG > 30 kJ/mol. Can we still use ATP? If so how?

How can a cell re-make spent ATP?

Couple it to an exothermic reaction of with DG < -30Longboard Create is in multiple steps similar to the generation of tRNA in the previous slide. Increase the concentration of ADP and Pi to force the reaction forward even if the DG0 is positive Substrate level phosphorylation

Other energy currencies in cells67

67The Currencies of energyATP

Other Phosphorylated compounds

Coupled reactions

Theioesters and Aceytl-CoA

Announcements: Change in syllabus: Please check it out on TED. Midterm now in on Mon Apr 27th.

Office hours: Friday 10-11am.

Homework was posted on Monday. Due in discussion section next week.

Slides will be posted today after lectures. Oxidation reduction Oxidation state for carbon

The redox currencies NAD+ and FAD

Thermodynamics of redox reactions

In groups of 2-3, sort all the molecules by the oxidation number of the marked carbon

Oxidation of Carbon atoms is exothermic2 C8H18+ 25 O2 16 CO2+ 18 H2O42.4kJ/ g

Reduction of NAD+ to NADH7373

Flavin Adenine Dinucleotide (FAD)7474

Reduction of FAD to FADH27575

Reduction of FAD to Semiquinone7676

Reduction of Semiquinone to FADH27777

Electrochemical Cell787879Electrons passed

from electron donor (reductant/reducing agent)

to electron acceptor (oxidant/oxidizing agent)1/2 O2 + NADH + H+ H2O + NAD+Half-reactionsNADH NAD+ + H+ + 2 e-electron donor1/2 O2 + 2 H+ + 2 e- H2Oelectron acceptorRedox Reactions80DG = nF DEn, number of electronsF, faraday (96.48 kJmol-1V-1 or 23.06 kcalmol-1V-1)DE, electrical potential difference or electromotive force or reduction potential (volts)How do we know which way the reaction goes?we need the Reduction PotentialFor spontaneous process, DG is negative, and so DE has to be positiveAnd like standard free energy DG, there is a standard reduction potential, DE 8081 DE = Ereduction + Eoxidation1/2 O2 + NADH + H+ H2O + NAD+NADH NAD+ + H+ + 2 e-electron donorEox = +0.315 V

So, back to our example: NADH reducing O2How do you get this number? Look in a big table.81

Standard Reduction PotentialsChange sign, because in our example, reaction is reverse NADH is being oxidized828283 DE = Ereduction + Eoxidation1/2 O2 + NADH + H+ H2O + NAD+NADH NAD+ + H+ + 2 e-electron donorEox = +.315 V

1/2 O2 + 2 H+ + 2 e- H2Oelectron acceptor Ered = +.815 VSo, back to NADH reducing O283

84This is the right sign oxygen is being reduced8485 DE = Ereduction + Eoxidation1/2 O2 + NADH + H+ H2O + NAD+NADH NAD+ + H+ + 2 e-electron donorEox = +.315 V

1/2 O2 + 2 H+ + 2 e- H2Oelectron acceptor Ered = +.815 VDE = .815 V + .315 V = 1.130 VCoupled reactions, add the partswere going to see this again when we cover oxidative phosphorylation85Oxidation reduction Oxidation state for carbon

The redox currencies NAD+ and FAD

Thermodynamics of redox reactions

Classification of Enzymes get a name based on what they do Oxidoreductase: Dehydrogenase, OxygenaseLyases: Aldolase, Decarboxylase Hydrolases: Phosphatase, ProteaseIsomerases: MutaseTransferases: Kinase, Ligases: CarboxylaseTypes of enzymes: IsomeraseConvert a molecule from oneisomer to another. Isomerases can either facilitate intramolecular rearrangements in which bonds are broken and formed.

Types of enzymes: KinaseCatalyzes the transfer ofphosphategroups fromhigh-energy, phosphate-donating molecules (often ATP) to specificsubstratesTypes of enzymes: Lyasecatalyzes the breaking of variouschemical bondsby means other hanhydrolysisandoxidation, often forming a newdouble bondor a new ring structure. Types of enzymes: DehydrogenaseOxidizes a substrate by a reduction reaction that transfers one or more hydrides (H) to an electron acceptor, usually NAD+/NADP+or a flavin coenzyme such as FAD or FMN LyaseTypes of enzymes: MutaseA type of isomerase that catalyzes the shifting of a functional group from one position to another within the same moleculeNote: direction of reaction that an enzymes could be named by is arbitrary

Answer to the GlycolysisPuzzle:94

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Introduction to Metabolism