Biochemical basis for therapy: Receptors and signalling

Question 1

Receptors are the sensing elements of chemical communication within the body being targets of what?

Answer 1

• Neurotransmitters (e.g. acetylcholine)
• Hormoes (“endocrines” e.g. adrenaline, insulin, glucocorticoid and reproductive steroids).
• A myriad of other mediators (e.. peptide growth factors, chemokines and cytokines of the immune system).
• Many therapeutic agents that act as agonists, antagonists, or allosteric modulators of selected receptor types frequently mimic, enhance, reduce, or block the effects of an endogenous substance.

Question 2

What does the ligand selectivity of receptors allow them to do?

Answer 2

A receptor due to its ligand selectivity responds to one, or more of the signalling molecules in the chemically-rich extracellular, or intracellular, environment - by analogy the “address” (target), to which the “letter” (signal) is posted.

Question 3

many cells are exposed to an essentially common extracellular fluid (ECF), why do they respond differently?

Answer 3

Although many cells are exposed to an essentially common extracellular fluid (ECF), they respond differentially because of the particular receptor types that they express.

Question 4

What are some examples of chemical signalling over short and long distances?

Answer 4

Autocrine - cell signals to itself
Paracrine - Cell signals to it’s close neighbours
Endocrine - (The signalling molecule enters the circulation). Cell signals via molecules transported by the blood to target distant cells.

Question 5

ow are receptors classified?

Answer 5

Receptors are protein (or glycoprotein) macromolecules classified on the basis of structure and function.

Question 6

What groups are they classified into?

Answer 6

Ligand-gated ion channels (LGICs) - aka ionotropic receptors.
G protein-coupled receptors (GPCRs) - aka metabolic receptors.
Kinase-linked receptors - aka enzyme-linked receptors
Nuclear receptors

Question 7

What are Ligand-gate ion channels (LGICs) - aka ionotropic receptors?

Answer 7

Located at the plasma membrane, targeted by hydrophilic signalling molecules (e.g. “fast” neurotransmitters (Acetylcholine, amino acids and rarely amines)) - action on a millisecond time scale.

Question 8

What are G-protein coupled receptors (GPCRs) - aka metabotropic receptors?

Answer 8

Located at the plasma membrane, targeted by hydrophilic signalling molecules (e.g. “slow” neurotransmitters (acetylcholine, amino acids, amines, peptides), adrenaline and peptide hormones) - signal on a second time scale (nb. many neurotransmitters can act as both rapidly and slowly via LGICs and GPCRs, respectively).

Question 9

What are kinase-linked receptors - aka enzyme-linked receptors?

Answer 9

Located at the plasma membrane, targeted mainly by hydrophilic protein mediators (e.g. insulin, numerous growth factors, many signalling molecules of the immune system) - work on an hours time scale.

Question 10

What are Nuclear receptors?

Answer 10

Located intracellularly in the nucleus ( or cytoplasm), targeted mainly by hydrophobic signalling molecules (e.g. steroid hormones, thyroid hormone) - very slow action on an hour/day time scale.

(Despite the name, some nuclear receptors are located in the cytoplasm and move (translocate) to the nucleus following activation by their agonist molecule).

Question 11

What are ion channels?

Answer 11

They are transmembrane pores formed by glycoproteins that span the membrane to create an ion conducting pathway for selected ions.

Question 12

How are Ion channels normally regulated and what does it affect?

Answer 12

They are usually regulated by signals that cause the channel to cycle reversibly between a closed (non-conducting) state and open (conducting) conformation - known as gating.
When open, conduct selected ions passively down their electrochemical gradients at incredible rates frequently mediating very rapid electrical signals (e.g. action potentials, synaptic potentials).

Question 13

What may ion channels be gated by?

Answer 13

Chemical signals (ligand-gated ion channels; LGICs)
Transmembrane voltage (voltage-gated ion channels; VGICs)
Physical stimuli (including thermal and mechanical energy).

Question 14

What is an example of a ligand-gated ion channel (LGICs)?

Answer 14

The nicotine acetylcholine receptor of skeletal muscle and neurones is the classic example.

Question 15

What do Ligand-gated ion channels (LGICs) consist of and what do they allow to occur?

Answer 15

Consists of separate glycoprotein subunits that form a central, ion conducting channel.
Allow very rapid changes in the permeability of the membrane to certain ions.
Rapidly (millisecond time frame) after membrane potential.

Question 16

What is the simplified sequence of LGIC activation?

Answer 16

Agonist binds - causing a rapid conformational change.
Channel opens - permitting a conduction pathway for selected ions.
Ions flow - down their electrochemical gradient: for the example of the nicotine acetylcholine receptor, movement of Na+ in to the cell and simultaneously movement of K+ out of the cell overall causes membrane depolarization and excitation.

Question 17

What is the process of signalling via second messengers?

Answer 17

Most cell surface receptors (by type) signal via second messenger systems in which receptor activation modulates that activity of an effector that is generally an enzyme (but can be an ion channel).
- many receptors, known as G protein-coupled receptors (GPCRs), are linked to a cell-membrane-located effector(s) by intermediary G proteins.
Enzyme effectors may increase, or decrease, their rate of synthesis of second messenger molecules affecting the activity of targets within the cell.
Ion channel effectors cause changes in membrane electrical properties.

Question 18

What is the basic structure of G protein-Coupled receptors (GPCRs)

Answer 18

• Integral membrane protein
• Single polypeptide with extracellular NH2 and intracellular COOH termini.
• Contains seven transmembrane alpha-helical spans joined by 3 extracellular and 3 intracellular connecting loops.
• May sometimes function as dimers (i.e. 2 separate receptors together)

Question 19

What is the basic structure of a G protein?

Answer 19

• Peripheral membrane protein
• Consists of 3 polypeptide subunits (alpha, beta and gamma)
• Contains a guanine nucleotide binding site in the alpha subunit that can hold guanosine triphosphate (GTP) or guanosine diphosphate (GDP).
• Exist as multiple types (e.g. Gs, Gj, Gg/11 (s, j, and g/11 are all subscript)) named according to their alpha subunit type.

Question 20

How is a G protein activated?

Answer 20

G proteins are activated by agonist binding to the GPCR to which they preferentially couple (via interaction between intracellular elements of the GPCR and G protein alpha-subunit)

Question 21

What is the difference between the inactive and active G protein molecules?

Answer 21

In the inactive (non-signalling) state he guanine nucleotide binding site of the alpha-subunit is occupied by GDP.
In the active (signalling) state the guanine nucleotide binding site of the alpha-subunit is occupied by GTP and the (alpha) and (Betagamma) subunits separate (dissociate) from each other.

Question 22

What is the result of the activation of a G protein?

Answer 22

The activation of a GPCR by an agonist causes a conformational change that is transmitted to the G protein alpha-subunit which:

• releases GDP and allows GTP to bind in its place (guanine nucleotide exchange)
• Separates from the receptor and betagamma-dimer (subunit dissociation).
• Generates a free GTP-bound alpha-subunit and Betagamma-dimer, both of which are signalling units.

Question 23

What is the state of a G-protein coupled receptor if it is not signalling?

Answer 23

• The receptor is unoccupied
• The G protein alpha-subunit binds GDP
• The effector is not modulated

Question 24

What is the state of a G-protein coupled receptor if it is turning on the signal?

Answer 24

• Agonist activates the receptor
• G protein couples with receptor.
• GDP dissociates from, and GTP binds to, the alpha subunit (i.e. guanine nucleotide exchange)
• G protein dissociates into separate alpha and betagamma subunits.
• G protein alpha subunit combines with and modifies, activity of effector.
• Agonist may dissociate from the receptor, but signalling can persist (because the G protein and receptor are now physically separate).

Question 25

What is the state of the G protein-Coupled receptor when turning off the signal?

Answer 25

• The alpha subunit acts as an enzyme (a GTPase) to hydrolyse GTP to GDP and Pi. The signal is turned off (nb.GTP does NOT dissociate allowing GDP to rebind).
• The G protein alpha-subunit recombines with the Betagamma subunit completing the “G protein cycle”

Question 26

What is an example of signalling via receptor kinases and what is the process of this?

Answer 26

THe receptor for insulin provides an example of one form of such signalling
Unbound
Binding of insulin causes autophosphorylation of intracellular tyrosine residues.
Recruitment of multiple adapter proteins, notably IRS1, that are also tyrosine phosphorylated.

Question 27

What is the process of signalling via Nuclear receptors (Class 1)?

Answer 27

Steroid hormones are lipophilic molecules - enter cells by diffusion across the plasma membrane.
Within the cell, they combine with an intracellular receptor producing dissociation of inhibitory HSP proteins. In the case of the steroid (but not thyroid hormone) receptors, the inactive receptor is located in the cytoplasm.
The receptor steroid complex moves to the nucleus, forms a dimer and binds to hormone response elements in DNA.
The transcription of specific genes is either “switched-on” (transactivated) or “switched-off” (transrepressed) to alter mRNA levels and the rate of synthesis of mediator proteins.