1. Mechanism of hormone action

2. Hormones Three types Proteins Lipids Amino acid derivatives
Glycoproteins
Small pepstides
Large proteins
Lipids
Cholesterol derivatives
Eicosanoids
Amino acid derivatives

3. Hormones Receptors Innate by themselves Require mediation
Binding sites for a hormone
Very specific

4. Hormone receptors Two types
Transmembrane
Intracellular/nuclear
Proteins regardless of the type
Interaction between a hormone and a receptor
Initial step of hormone action

5. Transmembrane receptors
Protein hormones
Unable to pass through the plasma membrane
Size
Charges
Receptors must be located on the plasma membrane
Extracellular domain for interaction with hormone
Intracellular signaling system

6. Types of transmembrane receptors
Receptors with multiple transmembrane domains
Seven trans-membrane domain receptor
No intrinsic enzymatic activity (C-terminus)
Associated with intracellular proteins involved in signaling
G-proteins
Modification of extracellular domain (hormone binding site, N-terminus)
Glycosylation
Crucial for hormone binding

8. Trans-membrane domains (7)
Alpha-helix
Hydrophobic amino acids
Loops
Connect alpha helices
May be linked by disulfide bridges (extracellular loop 1 and 2)

9. Intracellular/cytoplasmic domain
Palmitoylation of some cysteine residues
Attachment of fatty acids
Fourth loop
Site for phosphorylation

10. General structure of seven trans-membrane receptor
Variations
Amino acid sequences
Variable length of N-terminus
Affects binding of ligand/hormone

12. Intracellular signaling
Generated when a hormone interacts with extracellular domain of the receptor
Conformational change within the trans-membrane helices
Exchange of GDP to GTP on the alpha-subunit of G-protein complex
Activation of Ga subunit
Dissociation of activated Ga from G-protein complex (bg)

14. Second messengers Cyclic nucleotides (cAMP and cGMP) cAMP
Widely used secondary messenger
Generated by adenyl cyclase
Activated by activated Ga subunit of G-protein complex
Activation of cyclic nucleotide-dependent protein kinases
Protein kinase A (cAMP)

15. Secondary messengers Amplification of hormonal signals
Binding of hormone to the receptor
Activation of adenyl cyclase by activated Ga
Activation of protein kinase A by cAMP
Rapid clearance and inactivation
Phosphodiesterases
Inhibited by methylxanthines (caffeine, theophylline, and theobromine)
Phosphoprotein phosphatases

17. How do we know that cAMP is a secondary messenger?
Changes in production of cAMP after hormonal treatment
Correlation between amount of cAMP being produced and cellular response to the hormone
Inhibition of phosphodiesterase activity
Presence of ligand but no effects
Treatment with cAMP analogues/agonists
Similar response to that of hormone

18. Types of G-protein complex
Ga subunit (20 different types)
Gs (stimulatory Ga)
Gi (inhibitory Ga)
Go (associated with orphan receptors in neurons)
Gt (transducin found in retina, activates cGMP-specific phosphodiesterases)
bg complex
4 or more

20. Identification of specific G-protein complex associated with particular receptor
Structurally similar to each other
Use of pertusis toxin (bacterial toxin)
Uncoupling of G-protein complex from the receptor
Gi is very susceptible

21. G-protein complex coupled with secondary messenger system other than cyclic nucleotides
Generated through phospholipid metabolism
Inositol triphosphate (IP3)
Diacylglycerol (DAG)
Arachidonic acid
Activation of phospholipase C (PLC) by activated Ga

22. IP3 DAG Water-soluble Binds to protein kinase C
Stimulate release of Ca
DAG
Binds to protein kinase C
Activated by elevated Ca

23. Medical importance
65 % of prescription drugs target G-protein coupled receptors
Variety of ligands

24. Other protein hormone receptors
Transmembrane receptors with intrinsic tyrosine kinase activity
Receptor tyrosine kinase
Receptors for insulin and many growth factors
Transmembrane receptors with associated tyrosine kinases
Cytokine receptors
Receptors for growth hormone and prolactin
No intrinsic kinase activity
Interaction between receptor and hormone causes recruitment and activation of tyrosine kinases associated with receptor

25. Receptor tyrosine kinase
Approximately 100 receptor tyrosine kinases in human
Highly conserved
Domains
Extracellular
Hormone binding site
Transmembrane
Intracellular/cytoplasmic
Tyrosine kinase activity

26. 16 subfamilies Based on extracellular domain
Variation on extracellular domain
Interaction with variety of factors
EGF, PDGF, and insulin

27. Activation of receptor
Dimerization
Dimeric ligand (two subunits)
Each subunit binds to a receptor
Two binding sites within a hormone
One hormone interacts with two receptors

28. Activation of receptor
Pre-existence as a dimer
Receptor is a dimer
Activated through interaction with ligand

29. Activation of receptor
Conformational changes in the kinase domain
Accessible to the substrate
Autophosphorylation of tyrosine residues (3 in insulin receptor)
Activation loop
Triggers conformational changes
ATP binding
Interaction with intracellular proteins
Phosphorylation of other proteins