Enzyme kinetics

1. KEY CONCEPT
1) ∆G is independent
2) ∆G says nothing about the rate!

2. WHY DO WE WANT TO
KNOW ABOUT ∆G?

3. KEY CONCEPT
The value ∆G can be used to predict:
1)
2)

4. ∆G = ∆G° + 0.616kcal/M • ln [B]/[A]
A <-----> B
• ∆G = Cellular Change in Free Energy
• ∆Go = Standard Free Energy Change
• 0.616 = Gas/temperature constant in kcal/M
• [A] = concentration of A in M
• [B] = concentration of B in M

5. SO, WE NEED A WAY TO
GET ∆Go

6. DO EXPERIMENT IN THE
LAB
Mix equal amounts of A and B in test tube under
STANDARD STATE CONDITIONS and let it go
to equilibrium
Then evaluate the concentration of A and B

7. 0 = ∆G° + 0.616kcal/M • ln [B]/[A]
∆G° = - 0.616kcal/M • ln [B]/[A]
THUS:
WHAT HAPPENS TO ∆G AT
EQUILIBRIUM?

8. AND:
At equilibrium [B]/[A] is defined as
the k equilibrium (Keq)
THEREFORE:
∆Go = - 0.616kcal/M • ln Keq

9. SUMMARY
• ∆G = ??
• ∆Go = ??
• Keq = ratio of product [B] to substrate [A]
– Under what condition?

11. KEY CONCEPT
The correlation between ∆G and ∆G°
depends on the concentration of substrate
and product in the BODY

12. LETS TRY A
CALCULATION TO SEE
HOW IT WORKS

13. Glucose <-----> Fructose
A <----------> B

14. At equilibrium, the ratio of FRU to
GLU is 0.0475 at pH 7 and 23oC
• What is Keq?
• What is ∆Go’?
• What is ∆G?
• What does this info tell us about the reaction?

15. WHAT IS Keq?
Keq = [B]/[A]
Keq = [FRU]/[GLU]
Keq = 0.0475

16. WHAT IS ∆Go?
∆G° = -0.616kcal/mol(lnKeq)
∆G° = - 0.616kcal /mol • (ln0.0475)
∆G° = +1.88kcal/mol

17. WHAT DOES THIS TELL US
ABOUT THE RATE IN THE
BODY?
HOW ABOUT UNDER
STANDARD STATE
CONDITIONS?

18. WHAT IS ∆G?
∆G = ∆Go + 0.616kcal • ln[B]/[A]
BUT:
What do we need???

19. So we check cells and find that:
GLU [A] = 2 x 10-4M
FRU [B] = 3 x 10-6M
SO:
∆G = ∆G° + 0.616ln[B]/[A]
∆G = +1.88kcal/mol + 0.616*ln[3 x 10-6]/[2 x 10-4]
∆G = +1.88kcal/mol + (-2.58)
∆G = -0.71kcal/mol

20. SO WHAT DOES THIS MEAN?
In the body the reaction in which GLU is
converted to FRU is favorable and will
liberate -0.71kcal/mol of free energy.
What is the spontaneity?

21. SUPER MAJOR
KEY CONCEPT
The interplay between those reactions and
compounds with high levels of free energy
and those without is the whole essence of
metabolism.

22. NOTES ABOUT ENZYMES
• DO NOT affect the equilibrium, accelerate the
attainment of equilibrium

23. ENERGY, COUPLING AND
BIOCHEMISTRY

24. SUPER MAJOR
KEY CONCEPT
ALL reactions have a ∆G and a reaction
that produces a high level of free energy
can be used to drive a reaction that is
unfavorable.

25. KEY CONCEPT
In biological systems, a spontaneous chemical
reaction that is not at equilibrium is capable
of providing energy as it attempts to attain
equilibrium.

26. CONSIDER THE REACTION:
A B C D
-1.36 +5.45 -6.82

27. Total ∆G is additive for this set of reactions
-1.36 + 5.45 + (-6.82) = -2.73
A B C D
-1.36 +5.45 -6.82

28. Coupling can involve
TWO INDEPENDENT PATHWAYS.
A to B will not occur unless coupled to C to D.
A B
C D
+4.0
-7.3
A + C B + D -3.3

29. KEY CONCEPT
The overall free energy change for a
chemically coupled reaction is equal to the
sum of the free energy changes of the
individual steps

30. KEY CONCEPT
A thermodynamically unfavorable reaction
can be driven by a favorable one!

31. SO UP TO NOW:
Defined free energy
Intrinsic energy within molecules
Obtained as reaction moves to equilibrium
Indicated where it was found
(Chemical bonds, etc)

32. WHAT IS THE FORM OF
FREE ENERGY
AND
HOW DO WE OBTAIN AND
USE FREE ENERGY?

33. REMEMBER?
A -------> B = ∆G (-)

34. QUESTION?
HOW DO WE COLLECT THE FREE
ENERGY FROM THE CHEMICAL
REACTIONS?

35. OXIDATION
C6H12O6 + O2 -----> H2O + CO2
A -------> B

36. Oxidation of fuel molecules
generates free energy to drive
reactions
Glucose, Fatty Acids, Amino Acids

37. KEY CONCEPT
What happens in oxidation?
We need way to collect and transport this energy

38. Electron Carriers
• NAD +
• FADH +
• NADP +

39. NADH and FADH2
Transfer electrons to the oxidative
phosphorylation enzymes in the mitochondria
The energy in the electrons are used to
produce a H+ gradient so that ATP can be
synthesized

40. NADPH
Is the major
electron
donor in
reductive
biosynthesis

41. SO WE COLLECT ELECTRONS
WITH NADH
Can we use these electrons directly?

42. KEY CONCEPT
The free energy of electrons are used to
generate ATP
OXIDATIVE PHOSPHORYLATION

43. KEY CONCEPT
Adenosine triphosphate (ATP)
is the universal currency of free energy
in biological systems
PHOSPHORYL TRANSFER
POTENTIAL

44. ∆G° for the hydrolysis of ATP is
-7.3kcal/mol
ATP + H2O <----> ADP + Pi + H+

45. SO HOW DOES ATP WORK?
A B
ATP ADP + Pi
+ 4.0
-7.3
A + ATP B + ADP + Pi -3.3

46. 2-59 MBC

47. KEY CONCEPT
The hydrolysis of ATP changes the Keq by a
very large number and allows
thermodynamically unfavorable reactions to
occur.

48. ATP HYDROLYSIS
EQUALS AN INPUT OF ENERGY INTO A
REACTION
What will happen to equilibrium?

49. KEY CONCEPT
• A to B DO NOT have to represent a
chemical reaction!!!!
– Muscle contraction (movement of proteins)
– Ion flux (gradient)
– Change in Protein Conformation

50. OTHER COMPOUNDS ALSO
HAVE HIGH PHOSPHORYL
TRANSFER POTENTIAL
• Phosphenolpyruvate (PEP) ∆Go -14.3
• Creatine Phosphate ∆Go -10.3
• ATP ∆Go -7.3
• Glucose-1-phosphate ∆Go -5.3
• Glucose-6-phosphate ∆Go -3.3

51. THERE ARE MANY OTHER
ACTIVATED CARRIERS

52. COENZYME A
Universal carrier of acyl
groups
Carries 2-24 carbon units
Terminal sulfhydral group is a
reactive site

53. 2-62
MBC

54. SUMMARY
WE COLLECT THE FREE
ENERGY (∆G) IN TWO BASIC
FORMS
Electrons (e-)
High Energy Phosphate (PO4)

55. OK
You should know the following concepts:
∆G and its meaning
How enzymes fit into the mix
The form in which free energy is harvested
How free energy is utilized in the form of
ATP.