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Thermodynamics & ATP. Review thermodynamics, energetics, chemical sense, and role of ATP. Lecture 24. Thermodynamics in Biology. A Simple Thought Experiment. Driving Forces for Natural Processes. Enthalpy Tendency toward lowest energy state Form stablest bonds Entropy - PowerPoint PPT Presentation
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Thermodynamics & ATP
Review thermodynamics, energetics, chemical
sense, and role of ATP
Lecture 24
Thermodynamics in Biology
A Simple Thought Experiment
1 E. coli cell (10-11 mL)
1 mL H2O5 mg glucose1 mg (NH4)SO4
Mg++, PO4=, Fe3+, etc...
48 hours
109 cells1 mL H2O0 mg glucose<1 mg (NH4)SO4CO2
Glucose + (NH4)SO4 Cells + CO2
Driving Forces for Natural Processes
• Enthalpy– Tendency toward lowest energy state
• Form stablest bonds
• Entropy– Tendency to maximize randomness
Enthalpy and Bond Strength• Enthalpy = ∆H = heat change at constant pressure
• Units– cal/mole or joule/mole
• 1 cal = 4.18 joule
• Sign– ∆H is negative for a reaction that liberates heat
Entropy and Randomness
153 freeamino acids
Decreasedrandomness
Myoglobin
Entropy and Randomness
• Entropy = S = measure of randomness– cal/deg·mole
• T∆S = change of randomness
• For increased randomness, sign is “+”
“System” Definition
System
Surroundings
Closed system:No exchange ofmass or energy
“System” Definition
Isolated system:Energy is exchanged
E
E
“System” Definition
E
EM
M
Open System:Mass and energyare exchanged
Cells and Organisms: Open Systems
• Material exchange with surroundings– Fuels and nutrients in (glucose)– By-products out (CO2)
• Energy exchange– Heat release (fermentation)– Light release (fireflies)– Light absorption (plants)
1st Law of Thermodynamics
• Energy is conserved, but transduction is allowed
• TransductionOne form
of EAnother form
of E
Light Plants Chemicalbonds
Mayer: 1842
2nd Law of Thermodynamics
• In all spontaneous processes, total entropy of the universe increases
2nd Law of Thermodynamics
• ∆Ssystem + ∆Ssurroundings = ∆Suniverse > 0
• A cell (system) can decrease in entropy only if a greater increase in entropy occurs in surroundings
• C6H12O6 + 6O2 6CO2 + 6H2O complex simple
Entropy: A More Rigorous Definition
• From statistical mechanics:– S = k lnW
• k = Boltzmann constant = 1.3810–23 J/K• W = number of ways to arrange the system
• S = 0 at absolute zero (-273ºC)
Gibbs Free Energy• Unifies 1st and 2nd laws• ∆G
– Gibbs free energy– Useful work available in a process
• ∆G = ∆H – T∆S– ∆H from 1st law
• Kind and number of bonds– T∆S from 2nd law
• Order of the system
∆G• Driving force on a reaction• Work available distance from equilibrium• ∆G = ∆H – T∆S
– State functions• Particular reaction• T• P• Concentration (activity) of reactants and products
Equilibrium• ∆G = ∆H – T∆S = 0
• So ∆H = T∆S– ∆H is measurement of enthalpy– T∆S is measurement of entropy
• Enthalpy and entropy are exactly balanced at equilibrium
Effects of ∆H and ∆S on ∆G
Voet, Voet, and Pratt. Fundamentals of Biochemistry. 1999.
Standard State and ∆Gº
• Arbitrary definition, like sea level• [Reactants] and [Products]
– 1 M or 1 atmos (activity)• T = 25ºC = 298K• P = 1 atmosphere• Standard free energy change = ∆Gº
Biochemical Conventions: ∆Gº
• Most reactions at pH 7 in H2O
• Simplify ∆Gº and Keq by defining [H+] = 10–7 M
• [H2O] = unity
• Biochemists use ∆Gº and Keq
Relationship of ∆G to ∆Gº
• ∆G is real and ∆Gº is standard• For A in solution
– GA = GA + RT ln[A]• For reaction aA + bB cC + dD
– ∆G = ∆Gº + RT ln
– Constant Variable (from table)
º
[C]c [D]d
[A]a [B]b
}
Relationship Between ∆Gº and Keq
• ∆G = ∆Gº + RT ln
• At equilibrium, ∆G = 0, so
– ∆Gº = –RT ln
– ∆Gº = –RT ln Keq
[C]c [D]d
[A]a [B]b
[C]c [D]d
[A]a [B]b
Relationship Between Keq and ∆Gº
Keq G∆ º (kJ/m ol)10-6 34.310-5 28.510-4 21.410-3 17.210-2 11.310-1 5.91 0.0101 -5.9102 -11.3103 -17.2
Will Reaction Occur Spontaneously?
• When:– ∆G is negative, forward reaction tends to occur– ∆G is positive, back reaction tends to occur– ∆G is zero, system is at equilibrium
∆G = ∆Gº + RT ln [C]c [D]d
[A]a [B]b
A + B C + D
A Caution About ∆Gº
• Even when a reaction has a large, negative ∆Gº, it may not occur at a measurable rate
• Thermodynamics– Where is the equilibrium point?
• Kinetics– How fast is equilibrium approached?
• Enzymes change rate of reactions, but do not change Keq
∆Gº is Additive (State Function)
ReactionA BB C
Sum: A C
Also: B A
Free energy change∆G1º
∆G2º∆G1º + ∆G2º
– ∆G1º
Coupling Reactions
Glucose + HPO42– Glucose-6-P
ATP ADP + HPO42–
ATP + Glucose ADP + Glucose-6-P
∆Gºkcal/mol kJ/mol +3.3
+13.8 –7.3 –
30.5 –4.0 –
16.7
Resonance Forms of Pi
–
–
–
–
P
O
OHO
O
P
O
OHO
O
P
O
OHO
O
P
O
OHO
O
So: resonance stabilization
etc...
P
O
OO
O
Phosphate Esters and Anhydrides
ROH + HO P
O
O
O
R O P
O
O
OH2O
H2OEsters:
R CO
OH+ P
O
HO O
OH2O
H2O
R C
O
O P
O
O
O
Anhydrides:
= Hydrolysis
Hydrolysis of Glucose-6-Phosphate
∆Gº = –3.3 kcal/mol= –13.8 kJ/mol
O
CH2OP
O
O
HO
+ H2O O
CH2HO+ OHP
O
O
HOO
CH2OP
O
O
HO
+ H2O O
CH2HO+ OHP
O
O
HO
Ionization,resonance
Productstabilization
High ∆Gº Hydrolysis Compounds
∆Gº = –14.8 kcal/mol = –61.9 kJ/mol
OC
O
C
CH2
O P
O
O
O
Pi
H2OO
CO
C
CH2
OH
OC
O
C
CH3
O
Phospho-enolpyruvate
(PEP)
enol-ketoisomerization
High ∆Gº Hydrolysis Compounds
∆Gº = –11.8 kcal/mol = –49.3 kJ/mol
CO O
CH
PO O
O
HO
CH2 O P
O
O
O
1,3-Bisphosphoglyceric Acid(1,3-diPGA)
COO
CH OH
CH2OP
O
O
OH2O
Pi H
Resonancestabilized
High ∆Gº Hydrolysis Compounds
∆Gº = –10.3 kcal/mol = –43 kJ/mol
OC
O
CH2
N CH3
C NH
NH
P OO
O
H2O
OC
O
CH2
N CH3
C NH
NH2
+ Pi
PhosphocreatineResonancepossible
Phosphate Anhydrides (Pyrophosphates)
∆Gº = –7.3 kcal/mol = –30.5 kJ/mol
O P
O
O
O P
O
O
O P
O
O
O CH2O
Adenine
OHHORepulsionH2O
ADP + Pi
Adenosine triphosphate(ATP)
Thiol Esters
∆Gº = –7.5 kcal/mol = –31.4 kJ/mol
CoA S C
O
CH3
CoASH
+
+H3C C
O
O
H
H2O
Thiol Esters
R1C
SR2
O
R1C
SR2
O
R1C
OR2
O
R1C
OR2
O
Thiol ester less resonance-stabilized
“High-Energy” Compounds• Large ∆Gº hydrolysis
– Bond strain (electrostatic repulsion) in reactant• ATP
– Products stabilized by ionization• Acyl-P
– Products stabilized by isomerization• PEP
– Products stabilized by resonance• Creatine-P
“High-Energy” Compounds
• “High-energy” compound is one with a ∆Gº below –6 kcal/mol (–25 kJ/mol)
High-Energy Compounds
Group Transfer Potential
Lecture 25
Chemical Sense in Metabolism
Making and Breaking C–C Bonds
• Homolytic reactions
• Heterolytic reactions
A B A +B
A B A + B
Making and Breaking C–C Bonds
• Nucleophilic substitutions
R W + Z R Z + WLeavinggroup
Nucleophile
Nucleophilic Substitution Reactions
• SN1
a) R W R + WH
H W
b) R Z R Z+
Carbocation
CarbocationStability CH3 < 1o < 2o < 3o
Reactivity 3o > 2o > 1o > CH3
C R3R2
R1
(Planar)
Racemization
Common Biological Nucleophiles
R3
C W
R2R1
Z +
C
R3
WZ
R1 R2
R3
CZ
R2 R1
+ W
SN2 Nucleophilic Substitution
– –
Reactivity is SN2 ReactionsNucleophile ReactivityI–, HS–,RS– Very good >105Br–,HO–,RO–,CN– Good 104NH3;Cl–,RCOO– Fair 103; 101-102H2O, ROH Weak 1RCOOH Very weak 10-2
RO R CO
O>
Strongernucleophilic
base
Weakernucleophilicbase
Leaving Group
• Must accommodate a pair of electrons– And sometimes a negative charge
Major Role of Phosphorylation
• Converts a poor leaving group (–OH) into a good one (Pi, PPi)
PPi > Pi > H2O > OH
Acid Catalysis of Substitution Reactions
This H is often donated by an acidic sidechain of enzyme
ROH R OH+ Poorleaving group
ROH2 R + H2O Goodleaving group
ROHH
Central Importance of Carbonyls
1. Can produce a carbocation
2. Can stabilize a carbanion
C
O
C
O
C CO
C CO
Biological Carbonyls
R C
NH3
CO
O
H3C (CH2)n CO
O
R CH2 C
O
CH2 CO
O
COOC
O
CH2CH2OOC
Aminoacids
Fattyacids
β-keto acyl(fat oxiationan synthesis)
a-ketoglutarate(Kreβs cycle)
Aldol Condensation
H C
R1
R2
CO
R3
H
C
R1
R2
CO
R3
Aldol Condensation
C
R1
R2
CO
R3
O CR4
R5
O C
R4
R5
C
R1
R2
CO
R3
Aldol Condensation
O C
R4
R5
C
R1
R2
CO
R3
H
HO C
R4
R5
C
R1
R2
CO
R3
Aldolase Reaction• Glycolysis and gluconeogenesis
R4
CO
R5
+R1
CH
R2
CO
R3
Glyceraldehyde-3-phosphate
Dihydroxyacetone-phosphate
Aldolase
HO C
R4
R5
C
R1
R2
CO
R3
Fructose-1,6-bisphosphate
Claisen Condensationcarbanion + ester carbonyl ketone
R1 C
O
C
R5
R4
+
R2
C O
O
R3
R1 C
O
C
R5
R4
R2
C O
O
R3
Claisen Condensation
R1 C
O
C
R5
R4
R2
C O
O
R3
R1 C
O
C
R5
R4
R2
C O
+O R3H
HO R3
Thioesters in Biology
• In thioesters, the carbonyl carbon has more positive character than carbonyl carbon in oxygen ester.
R1 CO
O R2
R1 CO
S R2
Oxygen ester
Thioester
“High-Energy” Thioester Compounds
Coenzyme A
OAdenineCH2
OPOPO
O O
O O
CC
CH3
CH3
C
OH
H
C
O
NH
CCCO
HN
CH H
CH H
SH
β-mercapto-ethylamine
Pantothenic acid(growth factor)
ADP
O
P OO
O
OH
H
H
H
H
H
H
Fatty Acid Metabolism• Uses Claisen condensation
• Thiolase acts in fatty acid oxidation for energy production
SCoAC
CH2
O
R H2CC
O
SCoA
Thiolase
R CH2 C
O
CH2 C
O
SCoA + CoASH
Thiolase: Role of Cys-SH
Enz SH R C
O
S CoA
Enz S C
O
R CoASH+
+
R1 C
O
CH2
Enz SH + R C
O
CH2 C
O
R1
Thiolase: Role of Cys-SH
Enz SH I C C
H
H
O
O
+HI
Enz S C
H
H
C
O
O
Energy Diagram for Reaction
• ‡ is the transition state– Pentacovalent carbon, for example
Functional Groups on Enzymes
• Amino acid side chains–
–
– Imidazole–
SH
CO
O
CH2OH
Functional Groups on Enzymes
• Coenzymes/cofactors– Pyridoxal phosphate
• Metal ions and complexes– Mg2+, Mn2+, Co2+, Fe2+, Zn2+, Cu2+, Mo3+
Enzyme Inhibitors and Poisons
•
• Chelating agents– EDTA (divalent cations)– CN– (Fe2+)
• Cofactor analogs– Warfarin
• Suicide substrates
SH + Hg2+ S Hg(heavy metals)
Lecture 26
ATP and Phosphoryl Group Transfers
Phosphate Esters and Anhydrides
O P
OR
OH
O
O
PR2O OR1
ODiester
O P
OR
O
O
Monoester
Adenine ribose O P O P O P O
O O O
O O O
Mg2+Anhydride
Phosphoryl Group Transfers
OH
PHO
O
OH
OH
PHO OH
O
pK1=~1HO P
O
OH
O
pK2=6.7O P
O
OH
O
pK3=12.7
O
PO O
OP+ is electrophilePO4 hasnucleophilic O–
3-
Phosphoryl (Not Phosphate) Transfers
OCH2OH
OHOH
OHO P
O
O
O
HO
HO18-labeled(M. Cohn)
Glucose-1-phosphateO
CH2OH
OHOH
OHOH
P
O
O
OHO
+
Nucleophilic DisplacementsOR1
P O
OR3R2O
HOR4
OR1
P
OR4
O
R3O
R2O
OR1
P O
R4OR3O
+
R2OH
(nucleophile)
ATP as a Phophoryl Donor
• 2 roles for ATP– Thermodynamic
• Drive unfavorable reactions– Mechanistic
• Offer 3 electrophilic phosphorous atoms for nucleophilic attack
ATP as Phosphoryl Donor
• 3 points of nucleophilic attack
O P
O
O
O P
O
O
O P
O
O
O Ribose Adenine
aβg
Phosphor-ylation Pyrophos-
phorylationAdenylation (AMP)
Adenylyation: Attack on a-P
O P
O
O
O P
O
O
O P
O
O
O Ribose AdenineC CO
O
NH3
H3C
H
H3C C
H
NH3
C
O
O P
O
O
O Ribose AdenineO P
O
O
O P
O
O
O+
Alanine
Adenylation: Attack on a-P
H3C C
H
NH3
C
O
O P
O
O
O Ribose AdenineO P
O
O
O P
O
O
O+
PyrophosphatePPi
2Pi
Aminoacyl adenylate–Fatty acid activationfor oxidation-Amino acid activation for protein synthesis
Pyrophosphorylation: Attack on β-P
O P
O
O
O P
O
O
O P
O
O
O Ribose AdenineO
CH2P
O
O
O
OH OH
OH
AMP
P
O
O
O P
O
O
OCH2P
O
O
O
OH OH
O O
Ribose-5-phosphate
5'-phosphoribose-1-pyrophosphate(PRPP)
Phosphorylation: Attack on g-P
O P
O
O
O P
O
O
O P
O
O
O Ribose AdenineO
OHOH
OHHO
H2C OH
ADPGlucose
O
OHOH
OHHO
H2C O P
O
O
O
Glucose-6-phosphate
Amino Acid Sidechains as Nucleophiles
O P
O
O
O CH2
P-ser, thr
O P
O
O
NH
P-lys(e-amino)
N N P
O
O
O
P-his(1-N)
O P
O
O
NH C
NH2
NH
P-arginine
O P
O
O
NH C
NH2
NH CH2
P-creatine
COO
Enzymatic Phosphoryl Transfers
• Four classes– Phosphatases
• Water is acceptor/nucleophile– Phosphodiesterases
• Water is acceptor/nucleophile– Kinases
• Nucleophile is not water– Phosphorylases
• Phosphate is nucleophile
Phosphatases: Glucose-6-Phosphatase
O
OHOH
OHHO
CH2
O
P OO
OEnz X
O
OHOH
OHHO
CH2OH
Enz X P
O
O
OCovalent E-S intermediate is formed
X=His
Phosphatases: Glucose-6-Phosphate
Enz X P
O
O
O
HO
H
Enz X
P
O
O
O
HO
Phosphodiesterases: RNAase
O O
OHO
HHHH
PO
O
O
O
OHO
HHHH
PO
O
O
Pyr
Base
O O
OO
HHHH
PO O
Pyr
2',3'-cyclic phosphate
No covalent intermediate with enzyme
Phosphodiesterases: RNAase
O O
OO
HHHH
PO O
Pyr O O
OHO
HHHH
PO
O
Pyr
HO
H
OH
Kinases: g-Phosphoryl Transfer
• Transfer from ATP
O P
O
O P
O
O P
O
O Ribose Adenine
O O O
Mg2+
RX
OP
O
O
RX + O P
O
O P
O
O Ribose Adenine
O OMg2+
Kinases: P-Enzyme Intermediates
O P
O
O P
O
O P
O
O Ribose Adenine
O O OEnz X
ADP
OP
O
O
Enz X
Kinases: P-Enzyme Intermediates
OP
O
O
Enz X YNulceophilicsubstrate
Enz X O P
O
O
Y
Product
Kinases
RX Example Enz-X-P?R-OH Hexokinase
PFK?No
R-OP Nucleosidedisphosphokinase Yes
R-NH2 Creatine kinase NoR-COO– Succinate
thiokinase YesPyruvate kinase No
Protein-ser-OHProtein-thr-OH
Protein kinase YesR C
OCOO
Pyruvate Kinase• Makes ATP (∆Gº= –31 kJ/mol) from PEP
∆Gº= –62 kJ/mol
O P
O
O P
O
O P
O
O Ribose Adenine
O O OH2C CO
COO
H3C CCOOPyruvate
ADP +
O
PO O
H2C C
O
COOPEP
O
Phosphoryl-Group Transfer Potential
Compound ∆Gº StructurePEP -62 kJ/m ole Enol-P1,3-βisphospho-glycerate
-49 Acyl-P
P-creatine -43 Guani inium -PAcetyl-P -42 Acyl-PATP (an otherNTP)
-31 P-anhyrie
Glucose-1-P -21 Hem iacteal-PGlucose-6-P -14 Alcohol-PGlycerol-1-P -9 Alcohol
Significance of “High-Energy” P Compounds
• Drive synthesis of compounds below• Phosphated compounds are more
reactive– Thermodynamically– Kinetically
• If organism has ATP (etc…), it can do work and resist entropy
Cells must get ATP
