PEPTIDE HORMONES & HORMONES THAT USE CELL SURFACE RECEPTORS

1. PEPTIDE HORMONES& HORMONES THAT USECELL SURFACE RECEPTORS Endocrine mediators

2. OUTLINE: • Sizes, origins and fate. • Occurance in blood: bound/unbound. • Listing of major examples. • Receptors and receptor binding. • Tour of a selected peptide hormone: • insulin. • 6. Tour of a second peptide hormone:

3. Sizes, origins and fate: Due to the force of custom, “peptide hormones” is a collective name that has been applied to peptides, polypeptides and proteins that all function as hormones. The name “factor” has also been given to some of these peptides and originates from a time when their peptide/protein nature was still unknown. In the endocrine system of hormones originating from the brain, some of the smaller peptides that exist are from the hypothalamus and anterior and posterior pituitary (hypophysis).

4. SOME EXAMPLES OF SMALL PEPTIDES: THYROID RELEASING HORMONE (TRH) FROM THE HYPOTHALAMUS, A PEPTIDE OF ~360 daltons TARGET: ANTERIOR PITUITARY (RELEASE OF TSH) Glu-His-Pro-NH2 VASOPRESSIN FROM THE POSTERIOR PITUITARY, A PEPTIDE OF ~1040 daltons TARGET: KIDNEY & ARTERIOLES (INCREASES WATER RE-UPTAKE AND BLOOD PRESSURE)

5. OTHER PEPTIDE HORMONES FALL WITHIN THE DEFINITION OF A PROTEIN (>10 kD). SOME EXAMPLES: GROWTH HORMONE SOMATOTROPIN; 21.5 kD From the anterior pituitary; target: liver & bone CAUSES INCREASE IN METABOLISM & BONE GROWTH. This is a protein that has Considerable a-HELICAL AND RANDOM COIL STRUCTURE. THE BLUE ARROWS INDICATE APPROXIMATELY WHERE THE HORMONE BINDS TO CELL RECEPTORS.

6. PROLACTIN 22 kDwith a structure Somewhat similar to Growth hormone. Note the extensive a-HELICES. THE BINDING SITES FOR THE RECEPTORS ARE FROM THE LOWER SIDES. AS SHOWN BY THE BLUE ARROWS. THIS HORMONE IS MADE IN THE ANTERIOR PITUITARY TO STIMULATE MILK PRODUCTION.

7. FATE OF PEPTIDE HORMONES PEPTIDE HORMONES, AFTER BINDING TO THEIR RECEPTORS, ARE COMMONLY TAKEN UP BY THE CELLS THAT THEY BIND TO AND TRANSPORTED TO LYSOSOMES WHERE THEY ARE BROKEN DOWN . THE LYSOSOME, IS AN INTRACELLULAR SUBORGANELLE THAT CONTAINS SOME 30 DEGRADATIVE ENZYMES AS WELL AS AN ACIDIC pH ENVIRONMENT THAT FAVORS DESTRUCTION.

8. PEPTIDE HORMONESIN THE BLOOD • THE ABILITY OF A HORMONE TO REACH ITS TARGET • TISSUE IS CRUCIAL. THE FOLLOWING IS KNOWN ABOUT • THAT TASK: • SMALL PEPTIDE HORMONES ARE AT A LOW • CONCENTRATION AND SOLUBLE ENOUGH TO BE • TRANSPORTED UNBOUND TO ANY SOLUBLE CARRIER. • LARGE “PEPTIDE” HORMONES, LIKE GROWTH • HORMONE, ARE CARRIED BY A SOLUBLE PROTEIN • WHICH IS SIMILAR TO THE PROTEIN RECEPTOR • IT BINDS TO ON CELL SURFACES.

9. IMPORTANT PEPTIDE HORMONES(A SAMPLE LIST) Not part of the hypothalamic “pecking order” of hormones.

10. “TOUR” OF A SELECTED PEPTIDE HORMONE: INSULIN INSULIN IS A POLYPEPTIDE WITH A MW OF 5808. IT ACTUALLY CONSISTS OF 2 CHAINS JOINED BY DISULFIDE BONDS (A CHAIN = BLUE; B CHAIN = GREEN) IF YOU STRETCH OUT THE MOLECULE, YOU CAN SEE THAT THER ARE 3 DISULFIDE BONDS (1 INTRACHAIN ON THE A CHAIN AND 2 INTER- CHAIN BONDS)

11. SINCE ITS DISCOVERY IN 1921, INSULIN HAS BEEN ONE OF THE MOST HEAVILY STUDIED HORMONES OF ALL TIME. THIS IS DUE TO ITS ASSOCIATION WITH DIABETES, A DISEASE IN WHICH GLUCOSE BOTH STARVES SOME CELLS AND POISONSOTHER CELLS WHILE IT DISORGANIZES ASSOCIATED METABOLIC PATHWAYS. THE ROLE OF INSULIN IS TO FACILITATE THE UPTAKE OF GLUCOSE INTO CELLS BY MEANS OF GLUCOSE TRANSPORT PROTEINS. THAT PROCESS IS CONTROLLED BY TWO ASSOCIATED PEPTIDE HORMONES: GLUCAGON AND SOMATOSTATIN – ALL PRODUCTS OF THE PANCREAS.

12. INSULIN IS PRODUCED IN THE ISLET CELLS OF THE PANCREAS (B CELLS) GLUCAGON IS MADE IN SOME DIFFERENT ISLET CELLS CALLED A CELLS. SOMATOSTATIN IS MADE IN STILL OTHER ISLET CELLS CALLED D CELLS. THE ORGAN IS LOCATED BEHIND THE SPLEEN & ADJACENT TO THE SMALL INTESTINE.

13. INSULIN IS INITIALLY FORMED AS A SINGLE POLYPEPTIDE CALLED PREPROINSULIN. PROTEOLYTIC ENZYMES REMOVE THE SIGNAL PEPTIDE. THEN DISULFIDE BOND FORMATION TAKES PLACE TO FORM PROINSULIN. IN THE LAST STAGE, INSULIN IS FORMED WHEN CONTINUED LYSIS TAKES AWAY THE C PEPTIDE. THE HORMONE IS NOT ACTIVE PREVIOUS TO THIS STAGE.

14. BINDING OF INSULIN TO ITS RECEPTOR PROTEIN THE INSULIN RECEPTOR PROTEIN IS A TETRA- PEPTIDE COMPOSED OF TWO ALPHA & TWO BETA SUBUNITS. WHEN INSULIN BINDS TO THE PROTEIN, IT CAUSES A CONFORMA- TIONAL SHIFT THAT ACTIVATES KINASE ACTIVITY ON THE INSIDE OF THE MEMBRANE (ARROW). THIS PRODUCES A SERIES OF PHOSPHORYLATIONS AS A CASCADE OF MOLECULES (& FOLLOWING EVENTS). ONE EVENT IS THE DELIVERY OF GLUT4 TO THE CELL SURFACE.

15. POST RECEPTOR ACTIVITY: GLUT-4 SURFACE DELIVERY

16. SURFACE HORMONES THAT USE cAMP AS A MEDIATOR CORTICOTROPIN (ACTH) *** DOPAMINE EPINEPHRINE FOLLICLE STIMULATING HORMONE (FSH) *** GLUCAGON *** LUTEINIZING HORMONE (LH) *** MELANOCYTE STIMULATING HORMONE (MSH) *** PARATHYROID HORMONE *** PROSTAGLANDINS SEROTONIN SOMATOSTATIN *** THYROID STIMULATING HORMONE (TSH) *** *** peptides or polypeptides

17. GENERAL cAMPMECHANISM (stimulatory): THE KEY COMPONENTS ARE THE HORMONE (b-EFFECTOR); RECEPTOR (in membrane); THE G PROTEIN; ADENYLYL CYCLASE; AND cAMP. (cascade starts here)

18. An example of a hormone and its receptor: THE THYROID STIMULATING HORMONE AND THE TSHR PROTEIN THE MODEL SHOWS THE OUTERMOST PORTION OF THE RECEPTOR BOUND TO THE HORMONE ON THE LEFT AT THE LEUCINE RICH DOMAIN OF THE RECEPTOR. AS WITH ALL HORMONE-RECEPTOR BINDINGS, THE RECEPTOR WILL BE INDUCED TO AFFECT A NEARBY G PROTEIN PROBABLY IN THE VICINITY OF THE C-TERMINAL END OF THE RECEPTOR. IT IS CHARACTERISTIC OF THE HORMONE TO INDUCE A CHANGE IN THE CONFORMATION OF THE RECEPTOR THAT RESULTS IN SOME ACTIVITY OF THE RECEPTOR SUCH AS – ENZYME ACTIVATION ORPROTEIN BINDING.

19. WHEN A G PROTEIN IS AFFECTED, AT THIS POINT, THE G PROTEIN “PICKS UP” THE MESSAGE INITIATED BY THE HORMONE. TYPICALLY GDP (WHICH IS BOUND TO THE G PROTEIN) IS RELEASED AND REPLACED BY GTP AS A NEW ASSOCIATION WITH THE TRANSFORMED RECEPTOR PROTEIN. WHEN THIS OCCURS, THE g AND b SUBUNITS DIFFUSE AWAY FROM THE ORIGINAL G PROTEIN. AT 1, THE ACTIVATION BEGINS. BY 3, THE ACTIVATED PROTEIN DIFFUSES TO ADENYLYL CYCLASE TO ACTIVATE IT. IN STEP 4, THE G PROTEIN IS BACK TO ITS ORIGINAL INACTIVE FORM.

20. G PROTEINS COME IN SEVERAL VARIETIES (= ISOFORMS) TO INACT HORMONAL TRANSDUCTION MECHANISMS: NOTES: GS = STIMULATORY G PROTEIN SINCE IT STIMULATES ADENYLYL CYCLASE. GI = INHIBITORY G PROTEIN AS IT INHIBITS ADENYLYL CYCLASE. NOTE THAT OTHER TYPES ARE FOUND IN SPECIALIZED TISSUES SUCH AS BLOOD VESSELS, NOSE AND EYE.

21. AT THIS STAGE, GTP HAS BEEN FORMED AND HAS EITHER STIMULATED OR INHIBITED ADENYLYL CYCLASE. (cascade starts here) cAMP may be broken down at this point BEFORE it enters into any cascade. THIS IS ACCOMPLISHED WITH THE ENZYME cAMP PHOSPHO- DIESTERASE (PDE). HOWEVER, THE ENZYME CAN BE INHIBITED BY METHYL XANTHINES SUCH AS CAFFEINE AND THEOBROMINE (FOUND IN COFFEE, TEA, CHOCOLATE AND COLA DRINKS. THIS MAY RESULT, FOR EXAMPLE, IN INCREASED GI MOTILITY

22. CASCADE MECHANISMS: EXAMPLES OF THESE MECHANISMS HAVE BEEN SEEN BEFORE. A WELL- KNOWN ONE IS CONCERNED WITH THE BREAKDOWN OF GLYCOGEN INTO GLUCOSE WHEN THE HORMONE IS EPINEPHRINE. THAT’S SHOWN OFTEN SINCE MANY, MANY OF THE OTHERS HAVE NOT BEEN WORKED OUT WITH CERTAINTY – THOUGH EVIDENCE FOR THEIR EFFECTS ARE KNOWN. NOTE: IN PARTICULAR, THE ENZYME PROTEIN KINASE A (PKA) AND THE OVERALL AMPLIFICATION FROM x MOLECULES TO 10,000x MOLECULES IN THE CASCADE.

23. PROTEIN KINASES EXIST AT THE START OF MANY CASCADES AND EXIST IN A VARIETY OF ISOFORMS. THE ONE IN THIS CASCADE IS PROTEIN KINASE A (PKA) WHERE “A” IS SHORT FOR: cAMP. IN THE INACTIVE FORM (BEGINNING OF THE CASCADE), PKA HAS FOUR SUBUNITS WITH FOUR BINDING SITES FOR cAMP. WHEN cAMP BINDS TO THE SITES IT CAUSES THE C (OR CATALYTIC) SUBUNITS TO DISSOCIATE FROM THE R SUBUNITS AND FORM AN ACTIVE ENZYME. SHOWN ON THE RIGHT IS AN ACTIVATED SUBUNIT WITH ITS SUBSTRATE SHOWN IN RED. SOME OTHER PROTEIN KINASES ARE: PKB (USING A DERIVATIVE OF INOSITOL) AND PKC (USING CALCIUM)

24. THE LAST MECHANISM THAT WE WILL LOOK AT INVOLVES CALCIUM AND CALMODULIN THIS IS ACTUALLY A DOUBLE MECHANISM THAT REQUIRES TWO MEMBRANES, TWO ENZYMES AS WELL AS THE USUAL HORMONE, RECEPTOR PROTEIN AND A G PROTEIN (Gq) TO OPERATE. CALCIUM AND CALMODULIN PARTICIPATE IN PART OF THE MECHANISM. HORMONES ASSOCIATED WITH THIS MECHANISM INCLUDE PEPTIDE HORMONES SUCH AS VASOPRESSIN (POSTERIOR PITUITARY) AND THYROID RELEASING HORMONE (TRH). THIS IS A MECHANISM THAT MAKES USE OF PROTEIN KINASE C.

25. CALCIUM / CALMODULIN MECHANISM: CASCADE 1 CASCADE 2

26. SUMMARY • PEPTIDE HORMONES VARY FROM A FEW HUNDRED • TO THOUSANDS OF DALTONS IN SIZE. • MANY OF THEM ORIGINATE IN THE HYPOTHALMIC- • PITUITARY REGION. • EXCEPT FOR THE LARGEST HORMONES (PROTEINS) • THEY DO NOT REQUIRE CARRIERS IN THE BLOOD. • PEPTIDE HORMONES BIND TO RECEPTOR PROTEINS • ON THE CELL SURFACE. AFERWARDS, THEY ARE • INTERNALIZED AND DESTROYED. • 5. INSULIN IS A PEPTIDE HORMONE THAT IS MADE IN • THE PANCREAS TO TAKE GLUCOSE INTO SPECIFIED • INSULIN DEPENDENT CELLS. • 6. IT IS STORED IN INACTIVE FORM. WHEN RELEASED, IT • BINDS TO A RECEPTOR -- AN INACTIVE KINASE. • 7. THE KINASE STARTS A CASCADE OF EVENTS AMONG • WHICH IS GLUCOSE UPTAKE.

27. 8. WHEN cAMP (or cGMP) is used as a 2nd messenger, the process proceeds as follows: hormone > receptor protein > activation of G protein > activation of adenylyl (guanylyl) cyclase > cAMP (cGMP) > cascade. 9. CONTROL MECHANISMS ABOUND FOR THIS MECHANISM. 10. WHEN CALCIUM / CALMODULIN ARE USED AS 2ND MESSENGERS, THE MECHANISM PROCEEDS AS FOLLOWS: HORMONE > RECEPTOR PROTEIN > ACTIVATION OF G PROTEIN > ACTIVATION OF PHOSPHOLIPASE C TO RELEASE I tris P WHILE DAG ACTIVATES PKC ALONG WITH CALCIUM FREED BY I tris P. CALMODULIN OPERATES THE 2ND CASCADE WHILE CALCIUM (FREE) OPERATES THE 1ST CASCADE VIA PKC. WHEW!!