Signaling Pathways and Thermodynamics Recap 16

Recap of Signaling Pathways from Lecture 15

GPCR (G-Protein-Coupled Receptors)

  • Definition of GPCRs: G-protein-coupled receptors (GPCRs) are the largest and most diverse group of membrane receptors in eukaryotes.

  • Function: These cell surface receptors act like an inbox for messages, receiving signals in the form of:

    • Light energy

    • Peptides

    • Lipids

    • Sugars

    • Proteins

  • Role in Cells: GPCRs inform cells about the presence or absence of vital factors, such as nutrients or light, as well as conveying information from other cells.

  • Importance in Medicine: GPCRs play a critical role in numerous biological functions within the human body. Approximately one-third to one-half of all marketed drugs function by binding to GPCRs.

G Alpha Activation

Overview of Activation Process

  • GPCR structure includes:

    • G alpha (orange circles) interacting with GDP (guanosine diphosphate).

    • G beta-gamma (purple circles) coupled with the GPCR (light green loops).

Steps in Activation

  1. Agonist Binding: When a ligand, called an agonist, stimulates the GPCR, a conformational change occurs.

  2. Dissociation: G alpha dissociates from both the receptor and G beta-gamma.

  3. GDP to GTP Exchange: GDP is replaced by GTP (guanosine triphosphate), resulting in G alpha activation.

  4. Downstream Effects: Activated G alpha further activates other molecules within the cell.

G Proteins

  • Definition: G proteins are specialized proteins that can bind to the nucleotides GTP and GDP.

  • Structure: The G proteins associated with GPCRs are heterotrimeric, composed of three different subunits:

    • Alpha subunit

    • Beta subunit

    • Gamma subunit

  • Lipid Anchors: The alpha and gamma subunits are anchored to the plasma membrane via lipid modifications.

  • Comparison with Other G Proteins: Unlike heterotrimeric G proteins, some signaling proteins (like Ras) contain only a single subunit.

Hormonal Signaling Example: Epinephrine

Activation Steps in Response to Epinephrine

  1. Epinephrine Binding: Epinephrine binds to the GPCR (receptor).

  2. Conformational Change: This binding induces a conformational change in the receptor.

  3. G-Protein Activation: A conformational change in the G-protein occurs.

  4. Nucleotide Exchange: GDP is replaced by GTP on the G alpha subunit (Gα).

  5. Dissociation: Gα-GTP dissociates from G beta-gamma.

  6. Adenylyl Cyclase Activation: Gα activates adenylate cyclase.

  7. cAMP Production: Adenylate cyclase catalyzes the reaction of ATP to form cAMP (cyclic adenosine monophosphate) plus PPi (pyrophosphate).

cAMP and Glycogen Phosphorylase Activation

Mechanism of Action

  • Objective: How does cAMP activate glycogen phosphorylase?

  • Observation: The active form of glycogen phosphorylase is phosphorylated, while the inactive form is not.

  • Phosphorylation Process:

    • Phosphorylation of glycogen phosphorylase requires ATP.

    • The action is catalyzed by a kinase using phosphate from ATP.

    • Enzyme: Phosphorylase kinase catalyzes the phosphorylation of glycogen phosphorylase, converting it from the inactive (b) form to the active (a) form.

Role of Kinases in Phosphorylation

  • Function of Kinases: Kinases catalyze the addition of phosphate groups using phosphate from ATP.

  • Regulation: Phosphorylation is a common regulatory mechanism for enzymes, due to its speed and reversibility.

  • Chemical Reaction:

    • The general reaction can be shown as:
      extAminoacidwithahydroxylsidechain(serine,threonine,ortyrosine)+extATP<br>ightarrowextphosphorylatedaminoacid+extADPext{Amino acid with a hydroxyl side chain (serine, threonine, or tyrosine)} + ext{ATP} <br>ightarrow ext{phosphorylated amino acid} + ext{ADP}

Phosphorylate Phosphorylase Kinase and Further Cascades

  • Initial Phosphorylation: Phosphorylase kinase is activated by phosphorylation, changing from its inactive (b) form to the active (a) form.

  • Cascade Effect: Active phosphorylase kinase catalyzes the conversion of glycogen into glucose.

  • Role of PKA: cAMP activates protein kinase A (PKA), which in turn phosphorylates phosphorylase kinase.

  • Overall Process: Increases the overall cascade effect of the signal, magnifying the response.

Enzyme Cascades

  • Amplification: Enzyme cascades allow for massive signal amplification, as illustrated below:

    • 1 activated G protein → 20 activated adenylate cyclases → 100 active PKA → 1000 active phosphorylase kinases → 10000 active glycogen phosphorylase → resulting in substantial production of glucose.

Types of Cellular Responses

  • Responses Through Signaling: Various responses initiated by signaling pathways can include:

    • Opening of ion channels

    • Production of second messengers like cAMP

    • Direct activation of receptor enzymatic activities

    • Gene transcription in response to steroid hormones.

Growth Factors and Receptor Tyrosine Kinases (RTKs)

Signal Transduction Process

  1. Dimerization: Growth factors bind to two paired receptors, initiating a conformational change in the receptors.

  2. Kinase Activation: The receptors undergo autophosphorylation, activating their kinase activity.

  3. Ras Activation: Exchange of GDP for GTP occurs, activating Ras (a small GTPase).

  4. Enzyme Cascade Activation: The activation of downstream kinases (Raf, MEK, MAPK) in the pathway.

  5. Gene Transcription: Phosphorylated MAPK migrates into the nucleus to regulate gene expression involved in cell proliferation.

Steroid Hormone Receptor Signaling

  • Mechanism: Steroids can diffuse across cell membranes and bind to intracellular receptors.

  • Gene Regulation: Once bound, these receptors directly interact with DNA to regulate gene transcription, which is a slower, long-lasting, and generally irreversible process.

Characteristics of Signaling Pathways

  • Binding Properties: Ligands bind to their receptors with high specificity and affinity.

  • Involvement of Second Messengers: Signaling can involve the production of second messengers but not always.

  • Response Types: Cellular responses can vary in duration and reversibility; some are short-term while others are long-term and permanent.

  • Signal Amplification: The use of enzyme cascades is common, amplifying the initial signal significantly.

Basic Thermodynamics for Biologists: Laws of Thermodynamics

1st Law of Thermodynamics

  • Conservation of Energy: The energy of a closed system is conserved, although it can be converted from one form to another.

  • Entropy: You cannot decrease the entropy of a closed system.

2nd Law of Thermodynamics

  • Overview: Entropy of a closed system tends to increase over time.

Characteristics of Living Systems

  • Biological Composition: Living systems contain biological macromolecules (L-amino acids and D-monosaccharides).

  • Compartmentalization: They are compartmentalized, which facilitates internal regulation and system efficiency.

  • Functionality: Living systems extract energy from their environment to perform work and maintain homeostasis (defined as the maintenance of conditions suitable for life).

Importance of Energy Transduction

  • Ongoing Activity: Cells must continuously intake and convert energy into usable forms to maintain integrity and functionality.

  • Consequences of Neglect: Lack of energy transduction leads to cellular breakdown and loss of structural integrity.

Gibbs Free Energy (G and ΔG)

  • Total Usable Energy: Gibbs free energy is defined as the total usable energy within a system.

  • Spontaneity of Reactions:

    • If G_{end} < G_{start}, then energy is released (exergonic, favorable).

    • If G_{end} > G_{start}, then energy is consumed (endergonic, unfavorable).

Definitions of ΔG

  • Reaction Dynamics:

    • ΔG < 0: The reaction is exergonic and spontaneous.

    • ΔG > 0: The reaction is endergonic and non-spontaneous.

  • Free Energy Change Formula:

    • ΔG=Gproducts−GreactantsΔG = G_{products} - G_{reactants}

Entropy (S) and Change in Gibbs Free Energy

  • Entropy Change: Denoted as ΔS, where

    • ΔS=Sproducts−SreactantsΔS = S_{products} - S_{reactants}

  • Favorable Entropy Change: A favorable entropy change occurs when ΔS > 0.

Gibbs Free Energy Related to Enthalpy and Entropy

  • Relationship: Gibbs free energy is influenced by changes in enthalpy (ΔH) and entropy (ΔS) represented by the equation:

    • ΔG=ΔH−TΔSΔG = ΔH - TΔS

    • Where T is the temperature in Kelvin.

Standard Conditions for Gibbs Free Energy

  • Standard Conditions: Conditions for calculation include:

    • Temperature = 25°C

    • Pressure = 1 atm

    • Concentration of reactants and products at 1M

    • pH = 7.0

  • Equation:

    • ΔG°′=−RTextlnKeq′ΔG°' = -RT ext{ln} Keq',

    • Where R = gas constant, and T is temperature.

Calculating Equilibrium Constant (Keq) and Standard Gibbs Free Energy (ΔG°')

  1. Equilibrium Concentration Measurement:

    • Start with 1M of all reactants/products.

    • Let the reaction reach equilibrium and measure concentrations.

  2. Keq Calculation:

    • Keq′=rac[products][reactants]Keq' = rac{[products]}{[reactants]}

    • Determine if the reaction is exergonic or endergonic based on the value of Keq'. If

      • Keq' > 1: reaction is exergonic.

      • Keq' < 1: reaction is endergonic.

  3. Standard Gibbs Calculation:

    • ΔG°′=−2.303imesRimesTimesextlogKeq′ΔG°' = -2.303 imes R imes T imes ext{log} Keq'

Practical Example for Gibbs Free Energy

  • Example Scenario: Given equilibrium concentrations of A (1.95 M) and B (0.05 M), calculate:

    • Keq Calculation:
      Keq′=rac0.05M1.95MKeq' = rac{0.05 M}{1.95 M}

    • ΔG Calculation: Determining the sign of log can show whether the change is favorable or unfavorable.

ATP as a Primary Energy Source

  • Function of ATP: ATP is the primary source of energy in the cells, involved in many biochemical reactions.

  • Characteristics of ATP Hydrolysis: Hydrolysis of ATP is highly exergonic, releasing approximately -7.3 kcal/mol.

  • Energy Transfer: ATP hydrolysis is coupled with endergonic reactions to drive them forward.

Energy Coupling with ATP

  • Energy Transfer Methods:

    • Transfer of phosphate from ATP to substrates provides the energy needed for endergonic processes.

    • The overall reaction:
      C+ADP+Pi<br>ightarrowA+B+ATP+H2OC + ADP + Pi <br>ightarrow A + B + ATP + H2O

  • Creation of High-Energy Bonds: Newly created bonds in reactants exergonically dephosphorylate ATP, driving further reactions.

Conclusion

  • Summary of Thermodynamics and Energetics: Recap of:

    • Laws of thermodynamics

    • Gibbs free energy calculations for determining reaction spontaneity

    • Energy coupling and function of ATP in biological systems

  • Spontaneity Concerns: Highlighting that just because a reaction is termed spontaneous doesn't mean it occurs freely; context and reaction conditions are crucial.