Chapter 8.Chapter 8.

Metabolism & Metabolism & EnzymesEnzymes

Flow of energy Flow of energy through lifethrough life Life is built on chemical reactionsLife is built on chemical reactions

Chemical reactions of Chemical reactions of lifelife MetabolismMetabolism– forming bondsforming bonds between molecules between molecules

dehydration synthesisdehydration synthesis anabolic reactionsanabolic reactions

– breaking bondsbreaking bonds between molecules between molecules hydrolysishydrolysis catabolic reactionscatabolic reactions

Examples Examples dehydration synthesisdehydration synthesis

hydrolysishydrolysis

+

H2O

+

H2O

Examples Examples dehydration synthesisdehydration synthesis

hydrolysishydrolysis

Chemical reactions & Chemical reactions & energyenergy

Some chemical reactions Some chemical reactions release release energyenergy– exergonicexergonic– digesting polymersdigesting polymers– hydrolysis = catabolismhydrolysis = catabolism

Some chemical reactions require Some chemical reactions require input of energyinput of energy– endergonicendergonic– building polymers building polymers – dehydration synthesis = anabolismdehydration synthesis = anabolism

digesting molecules= less organization=lower energy

state

building molecules=

more organization=higher energy

state

Endergonic vs. exergonic Endergonic vs. exergonic reactionsreactions

exergonic endergonicenergy released energy invested

G

G = change in free energy = ability to do work

Energy & lifeEnergy & life Organisms require energy to live Organisms require energy to live

– where does that energy come from?where does that energy come from? coupling coupling exergonic reactionsexergonic reactions (releasing (releasing

energy) with energy) with endergonic reactionsendergonic reactions (needing (needing energy)energy)

+ + energy

+ energy+

Spontaneous Spontaneous reactions?reactions? If reactions are “downhill”, why If reactions are “downhill”, why

don’t they just happen don’t they just happen spontaneously?spontaneously?– because covalent bonds are stable because covalent bonds are stable Why don’t polymers

(carbohydrates, proteins & fats)

just spontaneously digest into their

monomers

Activation energyActivation energy Breaking down large molecules Breaking down large molecules

requires an initial input of requires an initial input of energyenergy– activation energyactivation energy– large biomolecules are stablelarge biomolecules are stable– must absorb energy to break bonds must absorb energy to break bonds

energycellulose CO2 + H2O + heat

Activation energyActivation energy the amount of energy needed to the amount of energy needed to

destabilize the bonds of a moleculedestabilize the bonds of a molecule– moves the reaction over an “energy hill”moves the reaction over an “energy hill”

Got a match? No, that’s too much energy

to get the work of life done!

Reducing Activation Reducing Activation energyenergy CatalystsCatalysts

– reducing the amount of energy to reducing the amount of energy to start a reactionstart a reaction

Pheew… that takes a lot less energy!

CatalystsCatalysts So what’s a cell to do to reduce So what’s a cell to do to reduce

activation energy?activation energy?– get help!get help! … chemical help… … chemical help…

Call in the...ENZYMES!

ENZYMES

G

Enzymes Enzymes Biological catalysts Biological catalysts

– proteins (& RNA) proteins (& RNA) – facilitate chemical reactionsfacilitate chemical reactions

increase rate of reaction without being consumedincrease rate of reaction without being consumed reduce activation energyreduce activation energy don’t change free energy (don’t change free energy (GG) released or ) released or

requiredrequired

– required for most biological reactionsrequired for most biological reactions– highly specifichighly specific

thousands of different enzymes in cellsthousands of different enzymes in cells

– control reactionscontrol reactions

Enzymes & substratesEnzymes & substratessubstratesubstrate

reactant which binds to enzymereactant which binds to enzyme enzyme-substrate complex: temporary associationenzyme-substrate complex: temporary association

productproduct end result of reactionend result of reaction

Enzymes & substratesEnzymes & substrates Enzyme + substrates Enzyme + substrates

productsproducts– sucrasesucrase

enzyme breaks enzyme breaks down sucrose down sucrose

binds to sucrose binds to sucrose & breaks disaccharide & breaks disaccharide into into fructose & glucosefructose & glucose

– DNA polymeraseDNA polymerase enzyme builds DNAenzyme builds DNA adds nucleotides to adds nucleotides to

a growing a growing DNA strandDNA strand

Lock and Key modelLock and Key model Simplistic model of enzyme Simplistic model of enzyme

actionaction– 3-D structure of enzyme 3-D structure of enzyme

fits substratefits substrate Active site Active site

– enzyme’s catalytic centerenzyme’s catalytic center– pocket or groove on pocket or groove on

surface of globular protein surface of globular protein – substrate fits into active sitesubstrate fits into active site

It’s shape that

matters!

Induced fit modelInduced fit model More accurate model of enzyme actionMore accurate model of enzyme action

– 3-D structure of enzyme fits substrate3-D structure of enzyme fits substrate– as substrate binds, enzyme changes as substrate binds, enzyme changes

shape leading to a tighter fit shape leading to a tighter fit ““conformational change”conformational change” bring chemical groups in position to catalyze bring chemical groups in position to catalyze

reactionreaction

How does it work?How does it work? Variety of mechanisms to lower Variety of mechanisms to lower

activation energy & speed up activation energy & speed up reactionreaction– active site orients substrates in active site orients substrates in

correct position for reactioncorrect position for reaction enzyme brings substrate closer togetherenzyme brings substrate closer together

– active site binds substrate & puts active site binds substrate & puts stress on bonds that must be broken, stress on bonds that must be broken, making it easier to separate making it easier to separate moleculesmolecules

Properties of Properties of EnzymesEnzymes

Specificity of enzymesSpecificity of enzymes Reaction Reaction specificspecific

– each enzyme is substrate-specificeach enzyme is substrate-specific due to fit between active site & substratedue to fit between active site & substrate

– substrates held in active site by weak interactionssubstrates held in active site by weak interactions H bondsH bonds ionic bondsionic bonds

– enzymes named for reaction they enzymes named for reaction they catalyzecatalyze sucrasesucrase breaks down sucrose breaks down sucrose proteasesproteases break down proteins break down proteins lipaseslipases break down lipids break down lipids DNA polymeraseDNA polymerase builds DNA builds DNA pepsinpepsin breaks down proteins (poly breaks down proteins (polypeptidespeptides))

ReusableReusable Not consumed in reactionNot consumed in reaction

– single enzyme molecule can catalyze single enzyme molecule can catalyze thousands or more reactions per thousands or more reactions per secondsecond

– enzymes unaffected by the reaction enzymes unaffected by the reaction

Factors that Affect Factors that Affect EnzymesEnzymes

Factors Affecting Factors Affecting EnzymesEnzymes

Enzyme concentrationEnzyme concentration Substrate concentrationSubstrate concentration Temperature Temperature pHpH SalinitySalinity ActivatorsActivators InhibitorsInhibitors

catalase

Enzyme concentrationEnzyme concentration

enzyme concentration

reac

tio

n r

ate

What’s happening here?!

Enzyme concentrationEnzyme concentration Effect on rates of enzyme activityEffect on rates of enzyme activity

– as as enzyme = enzyme = reaction rate reaction rate more enzymes = more frequently more enzymes = more frequently

collide with substrate collide with substrate

– reaction rate levels offreaction rate levels off substrate becomes limiting factorsubstrate becomes limiting factor not all enzyme molecules can find not all enzyme molecules can find

substratesubstrate

Substrate Substrate concentrationconcentration

substrate concentration

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n r

ate

What’s happening here?!

Substrate Substrate concentrationconcentration Effect on rates of enzyme activityEffect on rates of enzyme activity– as as substrate = substrate = reaction rate reaction rate

more substrate = more frequently more substrate = more frequently collide with enzymescollide with enzymes

– reaction rate levels offreaction rate levels off all enzymes have active site engagedall enzymes have active site engaged enzyme is enzyme is saturatedsaturated maximum rate of reaction maximum rate of reaction

37°

Temperature Temperature

temperature

reac

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ate

What’s happening here?!

TemperatureTemperature Effect on rates of enzyme activityEffect on rates of enzyme activity

– Optimum T° Optimum T° greatest number of molecular collisionsgreatest number of molecular collisions human enzymes = 35°- 40°C (body temp = 37°C)human enzymes = 35°- 40°C (body temp = 37°C)

– Increase beyond optimum T°Increase beyond optimum T° increased agitation of molecules disrupts bondsincreased agitation of molecules disrupts bonds

– H, ionic = weak bondsH, ionic = weak bonds denaturationdenaturation = lose 3D shape (3° structure) = lose 3D shape (3° structure)

– Decrease T°Decrease T° molecules move slower molecules move slower decrease collisionsdecrease collisions

Enzymes and Enzymes and temperaturetemperature Different enzymes functional in Different enzymes functional in

different organismsdifferent organisms

How do ectotherms do How do ectotherms do it?it?

7

pH pH

pH

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ate

20 1 3 4 5 6 8 9 10

pepsin trypsinWhat’s

happening here?!

pH pH Effect on rates of enzyme activityEffect on rates of enzyme activity

– protein shape (conformation) protein shape (conformation) attraction of charged amino acidsattraction of charged amino acids

– pH changespH changes changes charges (add or remove Hchanges charges (add or remove H++)) disrupt bonds, disrupt 3D shape disrupt bonds, disrupt 3D shape

affect 3° structureaffect 3° structure

– most human enzymes = pH 6-8most human enzymes = pH 6-8 depends on localized conditionsdepends on localized conditions pepsin (stomach) = pH 3pepsin (stomach) = pH 3 trypsin (small intestines) = pH 8trypsin (small intestines) = pH 8

Salinity Salinity

Salt concentration

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ate

What’s happening here?!

Salt concentrationSalt concentration Effect on rates of enzyme activity Effect on rates of enzyme activity

– protein shape (conformation) protein shape (conformation) depends on attraction of charged depends on attraction of charged

amino acidsamino acids

– salinity changessalinity changes change [inorganic ions]change [inorganic ions] changes charges (add + or –)changes charges (add + or –) disrupt bonds, disrupt 3D shape disrupt bonds, disrupt 3D shape

affect 3° structureaffect 3° structure

– enzymes intolerant of extreme enzymes intolerant of extreme salinitysalinity

Dead Sea is called dead for a reason! Dead Sea is called dead for a reason!

ActivatorsActivators Compounds which help Compounds which help

enzymesenzymes Cofactors Cofactors

– non-protein, small non-protein, small inorganicinorganic compounds & ionscompounds & ions Mg, K, Ca, Zn, Fe, CuMg, K, Ca, Zn, Fe, Cu bound in enzyme moleculebound in enzyme molecule

CoenzymesCoenzymes– non-protein, non-protein, organicorganic molecules molecules

bind temporarily or permanently tobind temporarily or permanently toenzyme near active siteenzyme near active site

– many vitaminsmany vitamins NAD (niacin; B3)NAD (niacin; B3) FAD (riboflavin; B2)FAD (riboflavin; B2) Coenzyme ACoenzyme A

Mg inchlorophyll

Fe inhemoglobin

InhibitorsInhibitors Regulation of enzyme activityRegulation of enzyme activity

– other molecules that affect enzyme other molecules that affect enzyme activityactivity

Selective inhibition & activationSelective inhibition & activation– competitive inhibitioncompetitive inhibition– noncompetitive inhibitionnoncompetitive inhibition– irreversible inhibitionirreversible inhibition– feedback inhibitionfeedback inhibition

Competitive Inhibitor Competitive Inhibitor EffectEffect

– inhibitor & substrate inhibitor & substrate “compete” for active site“compete” for active site

ex:ex: penicillin blocks enzyme penicillin blocks enzyme that bacteria use to build cell that bacteria use to build cell wallswalls

ex:ex: disulfiram (Antabuse) to disulfiram (Antabuse) to overcome alcoholismovercome alcoholism

ex:ex: methanol poisoning methanol poisoning

– overcome by increasing overcome by increasing substrate concentrationsubstrate concentration

saturate solution with saturate solution with substrate so it out-competes substrate so it out-competes inhibitor for active site on inhibitor for active site on enzymeenzyme

Non-Competitive Non-Competitive Inhibitor Inhibitor

EffectEffect– inhibitor binds to site other than active siteinhibitor binds to site other than active site

allostericallosteric site site calledcalled allostericallosteric inhibitorinhibitor

– ex: ex: somesome anti-cancer drugsanti-cancer drugsinhibit enzymes involved inhibit enzymes involved in synthesis of nucleotides in synthesis of nucleotides & therefore in building of DNA =& therefore in building of DNA =stop DNA production, stop DNA production, stop division of more cancer cellsstop division of more cancer cells

– ex:ex: heavy metal poisoning heavy metal poisoning– ex:ex: cyanide poisoning cyanide poisoning

causes enzyme to change shapecauses enzyme to change shape– conformational changeconformational change

renders active site unreceptiverenders active site unreceptive

Irreversible inhibitionIrreversible inhibition Inhibitor permanently binds to Inhibitor permanently binds to

enzymeenzyme– competitorcompetitor

permanently binds to active sitepermanently binds to active site

– allostericallosteric permanently changes shape of enzymepermanently changes shape of enzyme ex:ex: nerve gas, sarin, many insecticides nerve gas, sarin, many insecticides

(malathion, parathion…)(malathion, parathion…)– cholinesterase inhibitorscholinesterase inhibitors

doesn’t breakdown the neurotransmitter, doesn’t breakdown the neurotransmitter, acetylcholine acetylcholine

Action of Allosteric Action of Allosteric controlcontrol Inhibitors & activatorsInhibitors & activators

– regulatory molecules attach to regulatory molecules attach to allosteric site causing conformational allosteric site causing conformational (shape) change(shape) change

– inhibitor keeps enzyme in inactive forminhibitor keeps enzyme in inactive form– activator keeps enzyme in active formactivator keeps enzyme in active form

Cooperativity Cooperativity

Substrate acts as an activatorSubstrate acts as an activator– substrate causes conformational substrate causes conformational

change in enzymechange in enzyme induced fitinduced fit

– favors binding of substrate at 2favors binding of substrate at 2ndnd site site– makes enzyme more active & effectivemakes enzyme more active & effective

ex:ex: hemoglobin hemoglobin4 polypeptide chains: bind 4 O2; 1st O2 binds makes it easier for

other 3 O2 to bind

Metabolic pathwaysMetabolic pathways

AA BB CC DD EE FF GG enzyme

1

enzyme

2

enzyme3

enzyme4

enzyme5

enzyme6

Chemical reactions of life Chemical reactions of life are organized in pathwaysare organized in pathways– divide chemical reaction divide chemical reaction

into many small stepsinto many small steps efficiency efficiency control = regulationcontrol = regulation

AA BB CC DD EE FF GG enzyme

EfficiencyEfficiency Groups of enzymes organized Groups of enzymes organized

– if enzymes are embedded in membrane if enzymes are embedded in membrane they are arranged sequentiallythey are arranged sequentially

Link endergonic & exergonic reactionsLink endergonic & exergonic reactions

Whoa!all that going on in those

little mitochodria!

Feedback InhibitionFeedback Inhibition Regulation & coordination of productionRegulation & coordination of production

– product is used by next step in pathwayproduct is used by next step in pathway– final product is inhibitor of earlier stepfinal product is inhibitor of earlier step

allosteric inhibitor of earlier enzymeallosteric inhibitor of earlier enzyme feedback inhibitionfeedback inhibition

– no unnecessary accumulation of productno unnecessary accumulation of product

AA BB CC DD EE FF GG

allosteric inhibitor of enzyme 1

enzyme1

enzyme2

enzyme3

enzyme4

enzyme

5

enzyme6

X

ExampleExample– synthesis of synthesis of

amino acid, amino acid, isoleucineisoleucine from from amino acid, amino acid, threoninethreonine

Feedback inhibitionFeedback inhibition

Any Questions??Any Questions??