Lecture #5: Protein Structure and Function, Part 2

A set of practice flashcards covering key concepts from Lecture #5: Protein Structure and Function, Part 2.

What does Km represent in Michaelis-Menten kinetics?

The substrate concentration that yields half-maximal velocity; it reflects the enzyme–substrate affinity.

What does Vmax represent and what does it depend on?

The maximal reaction velocity at saturating substrate; depends on the amount of enzyme present.

What is activation energy (Ea) and why is it important?

The energy that must be overcome for a chemical reaction to occur; enzymes lower Ea to speed up reactions.

What is the transition state in a chemical reaction?

The high-energy state at the energy peak along the reaction path; reactants are converted to products at this point.

What is the difference between enzymes and inorganic catalysts?

Enzymes are biological catalysts (usually proteins, sometimes RNA in ribozymes) that are highly specific and often require cofactors/coenzymes; inorganic catalysts are non-biological and typically less substrate-specific.

What is an enzyme–substrate (ES) complex?

The intermediate formed when a substrate binds to an enzyme at the active site.

What is the difference between a substrate-binding pocket and a catalytic site?

The binding pocket binds the substrate; the catalytic site contains residues that facilitate bond making/breaking; they may overlap.

What is substrate specificity and how is affinity quantified?

Specificity is the ability to bind one molecule or group preferentially; affinity is the tightness of binding, often related to a low Kd (high affinity).

What is induced fit in enzyme catalysis?

Substrate binding induces conformational changes in the enzyme that improve alignment with the transition state.

Which substrates do trypsin, chymotrypsin, and elastase preferentially bind?

Trypsin binds positively charged Arg/Lys; chymotrypsin binds large hydrophobic residues; elastase binds small residues like glycine and alanine.

What is the Michaelis-Menten equation?

V0 = Vmax [S] / ([S] + Km); describes how reaction rate depends on substrate concentration for simple enzyme-catalyzed reactions.

How does pH affect enzyme activity?

Catalysis depends on the ionization state of catalytic residues; each enzyme has an optimal pH range due to ionizable groups.

How does temperature influence enzyme activity?

Increasing temperature raises kinetic energy and activity up to an optimum; excessively high temperatures can denature proteins.

What is competitive inhibition and its kinetic effect?

Inhibitor binds the active site, often resembling the substrate; Km increases while Vmax remains unchanged; can be overcome by high substrate concentration.

What is noncompetitive inhibition and its kinetic effect?

Inhibitor binds a site other than the active site; Vmax decreases while Km may remain unchanged; cannot be overcome by simply adding more substrate.

What are covalent modifications in protein regulation?

Covalent attachment of groups (e.g., phosphorylation, glycosylation, ubiquitination) that alter structure and activity.

What is allosteric regulation?

Non-covalent binding of a regulator at an allosteric site causes conformational changes that increase or decrease activity.

Describe calmodulin regulation by Ca2+ binding.

Calmodulin has four EF-hand Ca2+-binding sites; Ca2+ binding induces conformational changes allowing calmodulin to wrap around target sequences and alter activity.

What is the GTPase switch mechanism?

GTP-bound form is active; GDP-bound form is inactive; GEF promotes GDP→GTP exchange; GAP accelerates GTP hydrolysis to GDP.

What are kinases and phosphatases?

Kinases transfer phosphate from ATP to Ser/Thr/Tyr residues; phosphatases remove phosphate; together they regulate protein activity and localization.

What is phosphorylation and its role in regulation?

Addition of phosphate groups by kinases causes conformational changes and can activate or inhibit proteins.

What is glycosylation and its functional effects?

Attachment of sugar chains to amino acids; forms glycoproteins; affects solubility, targeting, folding, half-life, and interactions.

What is ubiquitination and proteasome-mediated proteolysis?

Attachment of ubiquitin chains to lysines; polyubiquitination marks proteins for degradation by the proteasome; monoubiquitination can alter function or localization.

How does the lysine used for inter-ubiquitin linkages affect function?

Different lysine residues in ubiquitin enable different linkage types; specific linkages determine substrate fate and activity; E3 ligases confer substrate- and linkage-specificity.

What are the three described forms of ubiquitination?

Polyubiquitination (degradation), monoubiquitination (altered function/localization), multiubiquitination (multiple single ubiquitins altering function/localization).

What is the advantage of assembling enzymes into multisubunit complexes?

Scaffold proteins reduce substrate diffusion time and increase pathway efficiency; some enzymes are fused into a single polypeptide to further minimize diffusion.

What are the main levels of protein regulation mentioned besides allostery?

Regulation can occur at synthesis/degradation (transcription/translation), covalent modifications, noncovalent interactions, and subcellular localization.

What is meant by the term 'binding affinity' in ligand interactions?

Affinity refers to the strength of the interaction; low Kd indicates high affinity, while high Kd indicates lower affinity.