Enzymes and the Rate-Limiting Step

• Spontaneous reactions can be slow and may require acceleration to sustain life.
• The rate-limiting step determines the overall reaction rate, and many uncatalyzed biological reactions proceed slowly.
• Increasing the number of molecules at the transition state increases the reaction rate.

Rate-Limiting Step

• Activation energy is required for molecules to interact and form products, but lower activation energy reduces the time needed.
• Enzymes lower activation energy, so they are not permanently altered and are required in small doses.
• Enzymes cannot affect reaction thermodynamics but are highly specific to substrates and regulated to meet cellular needs.
• Enzymes are proteins, except ribozymes, which are the exception.
• Most enzymes are proteins but may be conjugated with inorganic enzyme conjugates (cofactors) or organic enzyme conjugates (coenzymes).

Enzyme Components

• Prosthetic group: a tightly bound coenzyme or metal ion.
• Holoenzyme: a complete, catalytically active enzyme with its bound coenzyme and/or metal ions.
• Apoenzyme/apoprotein: the protein part of a holoenzyme.
• Active site: the specific environment where a reaction occurs rapidly.
• Substrate: the molecule bound to the active site.

Activation Energy (EA)

• A small energy input is required for any chemical transformation, but unlike free energy (ΔG), it varies by mechanism.
• Reaction molecules must reach a transition state to overcome it.

Ground State and Transition State

• Ground state: the starting point for forward or reverse reactions.
• Transition state (†): decay to substrate or product are equally likely.
• Biochemical standard free-energy change (ΔG'°)= standard change at pH 7.0.
• Activation energy (ΔG‡): the difference between ground state and transition state energy.
• Enzymes increase the number of substrate molecules with enough energy to reach the transition state.
• Activation energy and the number of molecules that reach the transition state change with reactions.
• The free energy of reaction remains the same.
• Enzymes affect reaction rates by decreasing activation energy, and a simple enzymatic reaction can be written as E + S ⇌ ES ⇌ EP ⇌ E + P.
• Enzymes catalyze the reaction of S to P (and P to S, under the right conditions), accelerate substrate interconversion, and are not consumed in the process.
• The equilibrium, reaction intermediate, and rate-limiting steps are simple enzymatic reactions.
• Reaction intermediate: any species on the reaction pathway with a finite chemical lifetime.
• Rate-limiting step: it determines the reaction's overall rate.

Enzyme Classifications

• Enzymes are grouped into seven classes that end in "-ase".
• Enzymes are named based on standard numbering systems by class, reaction type, and substrate.
• Isozymes are structurally different forms of an enzyme that catalyze the same reaction, and can have various tissue and developmental functions.
• Oxidoreductases: catalyze redox reactions, often using coenzymes like NAD+, NADP+, and FAD for electron transfer.
• Transferases: catalyze the transfer of chemical groups between molecules.
• Kinases transfer -PO42- between molecules.
• Amino transferases transfer amino groups, and DNA-dependent RNA and DNA polymerases are also transferases.
• Hydrolases catalyze hydrolysis by adding water to break bonds.
• Lyases catalyze the cleavage reactions not involving hydrolysis.
• Synthases (lyases) and synthetases (ligases) are enzymes with mechanisms that involve NTP intermediates.
• Isomerases catalyze molecular rearrangements to create isomers. Mutases move a phosphate group.
• Ligases catalyze joining carbons, typically energy from adenosine triphosphate (ATP)(synthetases) because it's from ATP hydrolysis).
• Translocases move ions/molecules across membranes.
• They create gradients, pump proteins into organelles, and may not alter molecule natures, and use energy and electrochemical conditions.

Active Site Specifics

• Enzyme + substrate forms enzymes-substrate complexes.
• Active site is the enzyme’s region for substrate binding with noncovalent bonds or transient covalent bonds.
• Enzymes have to be identical at the beginning and end.
• Coenzymes and cofactors required for catalysis.
• They May be covalently bound to the enzyme – prosthetic group.
• Accept electrons, temporarily re-orient substrate, bound in transition state, etc.
• Many, often come from vitamins, and ATP (IFF only energy, not a P used in product).
• Cofactors are generally metal ions.
• Enzymes without cofactors/coenzymes are called apoenzymes
• Everything, catalysis-related, are holoenzymes.
• Also true for transiently associated protein components
• Bacterial RNAP has σ (holoenzyme); without (apoenzyme)
• Coenzymes are often in the form of redox.
• Nicotinamide Adenine Dinucleotide (NAD) is one.
• It's in catabolic ones, but NADP-anabolic.
• C’OH ribose is attached to adenine.