Enzymes lower the activation energy of a reaction.
Substrate must have a matching shape to fit into the active site; two models describe enzyme binding to substrate.
Enzymes often require co-factors for activity.
Understand feedback inhibition and how cells regulate enzyme activity.
Cell Metabolism and Enzymes
Cell metabolism is organized by enzymes.
Chemical reactions in a cell would typically need much higher temperatures than exist within the cell.
Enzymes accelerate these reactions.
Reactions occur more slowly without enzymes.
Catalysts accelerate chemical reactions without being changed themselves.
Most enzymes are proteins (ribozymes).
Enzymes are highly specific due to their active sites.
Structure determines the specificity of the substrate and reaction.
Energy, Catalysts, and Metabolism
Catalysis allows the cell to control metabolism.
Catabolism: breaking down foodstuff into smaller molecules, generates energy.
Anabolism: uses energy to drive the synthesis of molecules.
Together, catabolism and anabolism constitute metabolism.
Enzymes can only catalyze reactions that would occur spontaneously.
Activation Energy and Free Energy
Enzymes lower the activation-energy barriers that block chemical reactions but do not change free energy.
Energy can never be created or destroyed; free energy is lost (energy that drives reactions).
Free energy is represented as ΔG.
A spontaneous reaction leads to a loss of free energy and occurs only if ΔG is negative.
Activation energy is required to “kick start” the reaction.
Lowering activation energy increases the probability of a reaction.
Activation Energy Details
Substrates pass through a series of intermediate states.
Energy is required to attain the most unstable intermediate state (transition state).
Input of activation energy is needed to reach the transition state.
Transition state: no longer a substrate, but not quite a product.
The transition state is a major determinant of reaction rate.
Reaction Rate and Equilibrium
Enzymes only alter the reaction rate but not the reaction equilibrium.
Enzymes decrease activation energy but do not change free energy.
Enzymes facilitate the formation of the transition state:
E+S↔ES↔ES‡↔EP↔E+P
Binding isn’t all or nothing; it involves association (K<em>on) and dissociation (K</em>off) rates.
Enzyme Reactions and Directionality
Enzyme reactions typically proceed as follows:
E+S↔ES↔E+P
The product is often thermodynamically stable and eventually reaches equilibrium.
Assuming irreversibility:
E+S↔ES→E+P
Reaction Pathways
Enzymes drive substrate molecules along specific reaction pathways.
Enzymes do not change the equilibrium point.
Enzymes lower the activation energy in both forward and reverse directions.
Enzymes do not change the direction of the reaction.
Substrate Binding
Substrates must fit the active site.
Enzyme shape determines whether a substrate will fit into the active site.
Specificity: binding of one molecule in preference to all others.
Enzyme Structure
All enzymes have:
Substrate binding site
Active site (catalytic domain)
Some have regulatory sites that change activity.
Regulatory sites are different from substrate binding sites.
Active Site Details
The active site is relatively small.
It's a 3D cleft made up of residues from multiple parts.
Non-polar microenvironment.
Substrate is bound by multiple weak non-covalent interactions.
Water is reactive and usually excluded.
Lock and Key Model
First model of enzyme-substrate interaction (1890s).
Substrate fits into a pre-existing site; must have matching shapes.
Specificity depends on the precisely defined arrangement of atoms in the active site.
Induced Fit Model
Second model of enzyme-substrate interaction.
Active sites are not necessarily rigid.
Shape can be modified by substrate binding.
Binding induces a conformational change, resulting in stronger binding.
The substrate plays a role in determining the final shape of the enzyme.
Binding of substrate to enzyme induces formation of active site.
Substrate strain induced by substrate binding to enzyme contorts the normal bonding angles and ‘activates’ the substrate.
Cofactors
Simple enzymes consist of protein only.
Some proteins require non-protein molecules (cofactors).
Cofactors provide chemical versatility not found in amino acids.
Protein + cofactor = complex enzymes or holoenzymes.
Apoenzyme (or apoprotein) = protein component.
Non-protein component = coenzyme or prosthetic group.
Cofactor Functions
Small non-protein molecules.
Functions:
Binds directly to substrate.
Allosteric change in enzyme shape.
Involved in transfer processes (e.g., NAD+, ATP).
Each cofactor associates with a variety of enzymes.
Types of Cofactors
~2/3 of enzymes require metals/trace elements as cofactors.
Some enzymes require several cofactors.
Classified based on:
Organic (e.g., heme, vitamin C, nucleotide (NAD)) or inorganic (e.g., metal ions (metalloenzymes) - trace elements in nutrition (Fe2+, Mg2+, Mn2+, Co2+, Zn2+, Cu2+, etc.)).
How they bind (coenzyme, prosthetic group).
Example: Alcohol Dehydrogenase requires zinc and NAD+.
Coenzymes vs. Prosthetic Groups
Prosthetic group:
Non-protein component tightly bound to apoenzyme by covalent bonds.
Coenzyme:
Non-protein component bound to apoenzyme non-covalently.
Many water-soluble vitamins act as coenzymes/prosthetic groups (e.g., Vitamin C, heme).
Clinical Symptoms of Dietary Enzyme Deficiency
Deficiencies affect various metabolic processes:
Niacin (nicotinic acid): Part of NAD; deficiency leads to pellagra.
B5 (pantothenic acid): Part of acetyl-CoA; deficiency leads to retarded growth, CNS disturbances.
B6 (pyridoxine): Coenzyme in amino acid and lipid metabolism; deficiency leads to retarded growth, anemia, convulsions, epithelial changes.
Folacin (folic acid): Coenzyme in amino acid and nucleic acid metabolism; deficiency leads to retarded growth, anemia, GI disorders, developmental abnormalities.
B12 (cobalamin): Coenzyme in nucleic acid metabolism; deficiency leads to impaired RBC production à pernicious anemia.
C (ascorbic acid): Coenzyme; delivers hydrogen ions, antioxidant; deficiency leads to epithelial and mucosal deterioration à scurvy.
Practice Exam Questions
Q: What is a non-protein molecule that associates with an enzyme to fully activate it? A. Cofactor
Q: What is the difference between a coenzyme and a prosthetic group? A. A coenzyme is bound to the apoenzyme non-covalently and a prosthetic group is bound covalently
Optimal Conditions
Every enzyme has an optimal temperature and pH.
Work over a limited pH range (e.g., pepsin = pH 2; trypsin = pH 7-9).
Outside this range can disturb ionic bonds and alter structure.
Temperature.
Too low, doesn’t work; too high = denature.
Increase in temperature increases energy collisions.
Enzyme Pathways
Enzymes often function in pathways.
Series of chemical reactions where products of one reaction become the substrate for the next reaction.
Regulation by Cells: Enzyme Production/Degradation
Increase/decrease the number of enzyme molecules depending on cellular requirements.
Done by either:
Altering transcription or translation.
Altering enzyme degradation by the proteasome.
Location
Enzymes need to be in the right location to perform the correct function.
Some enzymes have a fixed location (e.g., structural components of membranes, receptor tyrosine kinases involved in cell growth, lysosomal enzymes).
Substrate needs to reach a specific location for the enzyme to work.
Some enzymes can change location, e.g., CaMKII.
Activates transcription factors à learning.
Controls microtubules à cell division.
Post-Translational Modifications
Post-translational modifications (e.g., phosphorylation, glycosylation) can activate/inhibit some enzymes.
Alters activity by changing the shape.
These modifications are performed by enzymes (e.g., kinases, phosphatases, glycosyltransferases).
Zymogens
Enzymes synthesized as inactive precursors are called zymogens or pro-enzymes.
For full activity to occur, zymogens must undergo specific proteolytic cleavage (irreversible).
Blood clotting involves a series of zymogen activations of clotting factors.
Practice Exam Questions
Q: Which of the following is not a way in which cells regulate enzyme activity? A. Increasing the activation energy
Q: What are zymogens? A. Inactive enzyme precursors
Summary
Enzymes are catalysts; therefore, they remain unchanged.
Enzymes work by lowering activation energy but don’t change free energy.
Basic structure: substrate binding site, active site, some have regulatory sites.
Two models that describe substrate binding: Lock and key model; induced fit model.
Cofactors can increase enzyme activity.
Coenzymes and prosthetic groups.
Optimal conditions and factors influencing enzyme function (pH, temperature, ionic strength).
Cells can regulate enzymes in a variety of ways: feedback inhibition, altering the number of enzyme molecules, changing location, altering activity (post-translational modifications, proteolytic cleavage).
Further Reading
Alberts et al. Molecular Biology of the Cell. Sixth Edition. Chapters 2 and 3.
Alberts et al. Essential Cell Biology. Third Edition. Chapter 3.
Berg et al. Biochemistry. Sixth Edition. Chapter 8.