(Dr. Choy) (Unit A) Topic Note 5 Flashcards
What are the molecules that enzymes act on called?
Substrates
Are enzymes specific or general?
Specific, very specific
Do enzymes themseles get changed by the reaction?
No, they may change transiently (temporarily) during the reaction though
Do enzymes affect thermodynamics?
No, they do not affect the basic driving force behind chemcial transformations
What is the intermediate form between the initial chemical reactant and the product known as?
Transition state
What is the rate of the reaction controlled by?
Activation energy
How do enzymes affect reaction rate?
By lowering activation energy
Is the whole enzyme involved in catalysis?
No, only the active site of the enzyme is involved
Where are active sites usually found in an enzyme?
Between two domains or subunits
What factors contribute towards attaining the transition state?
- Bringing substrates together
- Orienting substrates in favorable geometry
- Supplying proton acceptors/donors, electron donors/acceptors
- Excluding water (sometimes)
- Stressing the substrate physically or electronically
- Selective binding of the transition state
What is chymotrypsin?
An enzyme secreted by the pancreas
* Allows breakdown of dietary protein during digestion
Chymotrypsin is a member of what family?
Serine protease
* Contains a critical serine residue in the active site
What does each serine protease contain?
A catalytic triad
* Serine, histidine, and aspartic acid
How does serine protease work?
Attack the peptide bond that joins amino acid residues to split the polypeptide in two at that site
What type of reaction do serine proteases carry out? Why is it called that?
Hydrolysis, because a molecule of water is consumed
How does chymotrypsin bind with substrate?
Binds such that the carbonyl carbon of the peptide bond is positioned for nucleophilic attack by the serine
What is the responsibility of the histidine in serine protease?
Acts as a general base to abstract a proton from the serine hydroxyl
What does the attack of the serine and histidine result in?
A tetrahedral intermediate
How does the tetrahedral intermediate break down?
The protonated histidine acts as a general acid to donate its proton to the nitrogen of the peptide bond
* Splits bond, releases carboxyl-terminal part of the peptide
When the peptide bond is split what happens to the serine?
Serine becomes covalently bound to the amino-terminal part of the peptide as an acyl-enzyme intermediate
What happens in the second phase of chymotrypsin activity?
Water enters
* Acts as nucleophile, activated by the histidine once again acting as the general base
What forms when water enters in chymotrypsin reaction?
Tetrahedral intermediate is formed
What happens to the chymotrypsin in the second phase of the chymotrypsin reaction?
Decomposes with histidine acting as a general acid such that the amino-terminal part of the peptide is released
What does the tetrahedral intermediate (transition state) contain? How is this managed?
An oxyanion
* Stabilized by entry into a special “hole” in the enzyme
How else is the transition state in chymotrypsin stabilized?
The h-bond between histidine and aspartic acid side hains becomes shorter and stronger
What strategies are used by chymotrypsin to lower transition states?
- Orientating substrates in a favorable geometry
- Supplying proton acceptors/donors
- Selective binding of the transition state
What are zymogens?
Enzymes synthesized that have a lack of enzymatic activity
What is the purpose of zymogens?
Allows the elaboration of enzyme activity to be precisely controlled
What is Vmax dependent on?
Enzyme concentration
* Doubling the amount of enzyme will double the rate of product formation
What does the Michaelis Constant represent?
Affinity of the enzyme for the substrate
* The lower the constant, the higher the affinity
Does enzyme concentration affect the Km?
No
Where is the Vmax on a Lineweaver-Burk plot?
The intercept on the 1/v0 axis
Where is Km on the Lineweaver-Burk Plot?
The intercept on the 1/[S] axis
What are the two classes of inhibitors?
- Irreversible: Forms covalent bond with enzyme
- Reversible: Binds non-covalently
What is a competitive inhibitor?
Bind directly in the active site and block substrate binding
* Many reversible inhibitors are this kind
What affect do competitive inhibitors have on Km and Vmax?
Increases Km without affecting Vmax
How can competitive inhibition be overcome?
High substrate concentration
How is the potent activity of enzymes controlled?
- Control of enzyme abundance (through gene expression)
- Synthesis of enzymes as inactive precursors (zymogens)
- Reversible modifications (phosphorylation etc.)
- Allosteric regulation
Where is allosteric regulation an important feature?
In metabolic pathways where precursor molecules are converted by a succession of enzymes into a product or products
