Enzyme Activity and Bioenergetics

1. Lower activation energy

1. Produce products in very high yields
2. Specificity determined by structure

1. Lyases

   Annotations:

   • Cleaves C--C, C--O, C--N or other bonds by elimination, leaving double bonds or rings, or addition of groups to double bonds
2. Isomerases

   Annotations:

   • Transfer of groups within molecules to yield isomeric forms
3. Ligases

   Annotations:

   • Formation of C--C, C--s, C--O, and C--N bonds by condensation reactions coupled to cleavage of ATP
4. Oxidoreductases

   Annotations:

   • Transfer electrons (hydride ions or H atoms)
5. Transferases

   Annotations:

   • Group transfer reactions
6. Hydrolases

   Annotations:

   • Hydrolysis reactions, transfers functional groups to water

1. Inorganics are essential ions or cofactors, organics are coenzymes
2. Apoenzyme is inactive form w/out cofactor, holoenzyme (is whole) is active with cofactor

1. Ask Woolridge
2. Enzymes preferentially bind to the transition state

1. Proton transfers are most common (10^2-10^5 rate enhancement)
2. AA residues can serve as catalysts
3. "Specific" involves H+ or OH- that diffuses into the catalytic center
4. "General" involves acids and bases other than H+ and OH-

1. Involves transient covalent bonds forming between enzyme and substrate
2. AAs that can do acid-base can do this as well

1. Mediate redox rxns
2. Stabilize charges
3. Ionize water

Annotations:

• ES is Michaelis complex of enzyme and substrate

Annotations:

• A constant derived from the rate constants. Under true Michaelis-Menten conditions is an estimate of the dissociation constant of E from S

1. SMALL Km MEANS TIGHT BINDING
2. BIG Km MEANS WEAK BINDING

Annotations:

• Constant at a given [E]

1. Theoretical max rate of the reaction but NEVER ACTUALLY REACHED (asymptotically approached as [S] increases)

Annotations:

• A turnover number, the number of substrate molecules converted to product/enzyme molecule/unit of time when E is saturated with S

1. k2 = kcat = Vmax/Et

Annotations:

• Measures how the enzyme performs when [S] is low

1. kcat/Km

1. x-intercept = -1/Km, y-intercept = 1/Vmax, slope = Km/Vmax

1. Can bind to and dissociate from the enzyme, often structural analogs and used as drugs
2. Noncovalent
   1. Competitive
      1. I binds only to E not to ES
      2. I and S bind at same site
      3. I does not affect catalysis
      4. Michaelis Menten Effects
         1. Km increases
         2. Vmax is unchanged
         3. Lines with different slopes but same y-intercept
   2. Uncompetitive
      1. I binds only to ES not to S
      2. I and S bind sites are different
      3. I does not affect substrate binding, but inhibits catalytic function
      4. Michaelis Menten Effects
         1. Km and Vmax decrease
         2. Ratio of Km to Vmax is same, so slope is same
         3. Parallel lines on Lineburker-Weaver plot
   3. Mixed
      1. I binds at E and ES
      2. I and S bind at different sites
      3. I binds enzyme with or without substrate
      4. Binding inhibits both substrate binding and catalysis
      5. Michaelis Menten Effects
         1. Vmax decreases
         2. Km can increase or decrease (unchanged in extreme cases of noncompetitive
         3. Wacky lines, x-intercept, y-intercept and slope are all different
      6. Becomes noncompetitive in extreme cases

1. Permanently shut off enzyme, often toxins but sometimes used as drugs
2. Covalent

1. Compartmentalized
2. Catabolic converge on acetyl-CoA
3. Anabolic diverge from few metabolites

1. PEP and 1,3-BPG are the highest
   1. PEP
      1. Largest free energy hydrolysis
      2. Hydrolysis yields enol form of pyruvate which tautomerizes to the keto form
   2. 1,3-BPG
      1. Mixed anhydrides or acetyl phosphates
      2. Bond strain, electrostatics, and resonance create high energy
2. ATP is mid level, good for transporting P groups
   1. Large negative free energy change on hydrolysis due to
      1. Relief of electrostatic repulsion
      2. Stabilization of products by ionization
      3. Stabilization of products by resonance
      4. Entropy factors
      5. Greater degree of hydration of prodcuts
3. Thioesters like acetyl CoA
   1. Orbital overlap between carbonyl and sulfur is not as good as resonance between oxygen and carbonyl in esters

1. Reduced compounds serve as fuel which electrons can be stripped off of during oxidation
2. Transfer of electrons is often accompanied by the transfer of protons and hydride
3. NAD and NADP are common redox cofactors
4. Flavin cofactors use molecular oxygen as ultimate electron acceptor, tightly bound to proteins and allow single electron transfer