Major concepts

1. What is biosignaling, why is it needed, and what are its characteristics?
2. For each biosignaling method we study, how does the receptor respond?
3. What makes a signal end? What is desensitization, and how does it occur?
4. How does a cell restore its pre-signal internal environment?
5. How can biosignaling pathways differ from each other?

Core knowledge

1. What are the domains of a receptor?
2. How does the nicotinic acetylcholine receptor function? What happens as a result?
How are the results reversed?
3. How does a 7tm receptor function?
4. What are the subunits of a G protein, and what does each do? How is a G protein activated, and how is it inactivated?
5. What reaction does adenylyl cyclase (AC) catalyze? How is its product removed from a cell?
6. What are the subunits of protein kinase A (PKA), and how is PKA activated?
7. What happens if a different G protein is activated?
8. For all three receptors (nicotinic ACh, 7tm, and insulin), which molecules are proteins and which are not? Which proteins are enzymes, and what reactions do the enzymes catalyze?

Note: The introduction to this chapter is an example of a significant improvement in the 4^th edition of the text compared to the 3^rd edition.
Also, this area of study is exploding: biosignaling is important in cell biology, developmental biology, cancer, immunology, neurophysiology, and more.

Biosignaling: chemical and electrical methods of communication → change in a cell
A. Characteristics (Lehninger)                                                                                               Fig. 12-1
     1. Specificity: a specific protein-ligand interaction →
               change in protein conformation and function
     2. Amplification: one enzyme modifies several others; each of those modifies several
     3. Desensitization/adaptation: feedback → reducing or turning off signal
     4. Integration: Opposing signals are integrated, and the cell responds to the sum.
B. Additional characteristics (Martin)
     1. Reversible: The protein-ligand complex dissociates, the phosphate is removed, etc.
     2. Sequence: A series of reactions/interactions occurs.
     3. Enzymes can be affected allosterically, covalently, or both.
         Allosteric regulation is not amplified; covalent regulation can be amplified.
     4. Versatility: One signaling pathway can affect multiple enzymes, leading to
              regulation of several metabolic pathways.
C. Types of systems (we'll study 1* and 3*)                                                                        Fig. 12-2
     1*. Gated ion channels open in response to a signal. Change in [ion] → cell change.
     2. Receptor enzyme is activated by ligand binding. Product of reaction → cell change.
     3*. Receptor protein is activated by ligand binding → indirect activation of an enzyme
               → cell change
     4. Nuclear receptor is activated by ligand binding → change in, eventually, protein
              synthesis → cell change
     5. Receptor protein directly activates enzymes → cell change
     6. Receptors → cytoskeleton changes → cell change

Gated ion channels
A. Background:
      1. Membrane potential is created and maintained by Na⁺–K⁺ ATPase.
      2. Ca²⁺ and Cl⁻ also have significant concentration gradients across the membrane.
      3. Opening an ion channel → spontaneous flow of ions across the membrane.
          Na⁺, for example, moves into the cell → depolarization of the membrane.
          The number of Na⁺ required to depolarize the membrane is <<<<the number required to change C₁ or C₂.
      4. The equilibrium potential (E) for an ion is when R T ln (C₂/C₁) = Z F ΔV.
B. Nicotinic acetylcholine receptor                                                                     Fig. 11-51, Fig. 12-4
     1. Acetylcholine (ACh) is the ligand = choline with an ester bond to acetic acid
         
Reaction is reversed by acetylcholinesterase.
          Receptor also binds nicotine:
     2. Receptor is a gated ion channel that allows Na⁺, Ca²⁺, or K⁺ to cross the membrane.
          Normally the channel is closed.
     3. Binding of 2 ACh in the 2 binding sites → conformation change with open channel.
     4. Channel closes after a short time, either because 2 ACh leave or as a result of
              desensitization. Mechanism of desensitization is unclear.
C. Voltage-gated ion channels open (or, in a few cases, close) in response to voltage.
     Types: Na⁺, K⁺, and Ca²⁺ channels
     Distribution: at different locations on neurons and muscle cells
D. Results of opening ion channels temporarily
     1. Transmission of an action potential (message) along a neuron
               by depolarizing, then repolarizing the membrane
     2. Transmission of an action potential from one neuron to another
              Ca²⁺ causes release of ACh into the synapse so that it can bind to the next neuron.
     3. Muscle cell contraction in response to increased [Ca²⁺], etc.
E. Other types of neuronal ion channels respond to other neurotransmitters.

Receptors that indirectly activate enzymes include receptors that activate G proteins and receptors that cause an increase in intracellular [Ca²⁺+ ]

Overview of the G protein system and sequence:
1. Hormone binds a receptor
2. Receptor then activates a G protein, which releases GDP and binds GTP
3. G protein activates an enzyme, which produces a 2^nd messenger
4. 2^nd messenger activates an enzyme that covalently modifies other enzymes.

Receptor: integral protein with seven transmembrane α-helices (7tm protein)
Example: β-adrenergic receptors (two types of receptors for epinephrine)
1. Studied using agonists (bind instead of epinephrine → same effect) and antagonists
      (prevent binding by epinephrine → no effect)
2. Effect: Receptor with ligand bound causes G protein to release GDP and bind GTP.
3. Amplification: One receptor can activate several G proteins.
4. Desensitization: phosphorylation by β-adrenergic protein kinase (βARK) leads to
      binding by β-arrestin, which prevents G protein binding by the receptor.

G protein
A. Structure
      1. Gαβγ inactive (heterotrimeric) has αβγ subunits; binds GDP stably; binds receptor
     2. G_sα active = α subunit only; binds GTP and hydrolyzes GTP to GDP + P_i
      3. G_sα is a lipid-anchored membrane protein.
B. Activation of G_sα requires receptor to initiate
     1. release of GDP
     2. binding GTP
     3. separation from Gβγ (which can also activate enzymes in the cell).
C. Activated G_sα binds and activates AC.
      Once G_sα converts GTP to GDP, the G protein is inactivated and leaves AC.
D. Types of G proteins: G_sα, G_iα, G_qα, several others

Adenylyl cyclase (AC)
A. Structure and location: integral membrane protein with a binding site for G_sα or G_iα .
B. Reaction: ATP → cAMP + PP_i   (occurs several times)
C. Active only with G_sα bound

cAMP = 2^nd messenger (1^st messenger = epinephrine)
A. synthesized by AC from ATP in an irreversible reaction
B. binds and activates protein kinase A (PKA) and other enzymes
C. converted to AMP by cyclic nucleotide phosphodiesterase (hydrolase)

Protein kinase A (PKA)
A. Structure
     1. Inactive form = R₂C₂ (2 regulatory subunits bound to 2 catalytic subunits)
         R subunits have pseudo-consensus sequence bound by active site of C subunits.
     2. cAMP binds to 2 allosteric sites on R subunits → conformation change,
         release of C subunits
B. Active PKA (each C subunit = 1 PKA) catalyzes transfer of P from ATP to S/T in consensus sequence
C. PKA is inactivated when C subunits are bound by R subunits.
     Phosphorylation of enzymes is reversed by protein phosphatases.

Desensitization of the pathway
A Phosphorylation of the receptor when it has had ligand bound for longer than usual
     by β-adrenergic receptor kinase (βARK)
B. Creates a binding site for β-arrestin, which prevents G protein binding
C. Reversed by removing the receptor from the membrane (endocytosis),
     followed by dephosphorylation, then returning the receptor to the membrane.

Variations on the G protein pathway
A. Other receptors can activate G_sα, which means other hormones have the same effect.
B. Other G proteins can be activated by receptors
     1. G_iα inhibits AC; G_iα is activated by α₂-adrenergic receptors, for example.
     2. G_qα activates phospholipase C, activating a different G protein pathway.
         Phospholipase C hydrolyzes phosphatidyl-inositol-4,5-bis-phosphate to
         a. IP₃ = a ligand that binds a receptor channel for Ca²⁺ → increased [Ca²⁺]_cytosol
         b. DAG = activates protein kinase C (PKC)
          IP₃, DAG, and Ca²⁺ are additional 2^nd messengers that cause changes in the cell.
C. cAMP can affect the activity of enzymes other than PKA.

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