Cyclic AMP and Hormone Action

1. Cyclic AMP and Hormone Action Hormones are important regulators of biological processes. Those that work through the action of 3’-5’-cyclic AMP (cAMP) control enzymes by using ATP to phosphorylate serine and threonine groups on target enzymes. These so-called protein kinases represent a sequel of catalytic steps designed to amplify the action of the hormone. The key word here is “catalytic”, which means many progeny can come from one. There is a pattern to many protein kinase-dependent processes and this tutorial will help you see commonality in the mechanism. As you study the steps, asks yourself why doesn’t the hormone just engage the enzyme directly. You will soon realize that hormones are present in fleeting amounts and cannot be effective at these minute levels. Rather their action depends on events that allow the meek hormone to raise to a higher level of effectiveness and deliver a mighty blow to the cell process in question. The meek to the mighty helps you see how this is possible. From the Meek to the Mighty

2. Hormone receptor Initial response Let’s begin by examining a typical c-AMP-dependent hormone response system. First we must realize that cAMP action is inside the cell and the hormone is outside. A connection, therefore, must be made across the membrane. The connecting point is a receptor for the hormone in close proximity to a membrane bound enzyme, adenylcyclase (click 1). Adenylcyclase When the hormone contacts its receptor it sets into motion a series of reactions that ultimately lead to the synthesis of cAMP (click 1). GDP GTP cAMP G protein ATP The first reaction is the exchange of GDP with GTP on G protein (click 1). This allows an “active” G protein to travel to adenylcyclase and stimulate the enzyme to make cAMP from ATP (click 1). Click 1 to go on.

3. cAMP C R cAMP R R + 4 cAMP + cAMP C C C R cAMP cAMP-dependent Protein Kinase The next step in the reaction sequence is to activate the first kinase enzyme, which is called cAMP-dependent protein kinase or cAPK for short. The catalytic subunits of cAPK are held in check by regulatory subunits (click 1). But, when cAMP is around, the regulatory subunits bind the cAMP and dissociate from the catalytic subunits (click 1). The catalytic subunit is now free to attack a protein target. You should note at this stage that by raising cAMP levels through an enzyme, the hormone has dramatically amplified its ability to elicit a response. This is the advantage of a “catalytic” (one begets many) as opposed to a stoichiometric (one begets one) response. Click 1 to go on.

4. cAPK 2ATP 2ADP         P P Calmodulin Inactive Phosphorylase Kinase Active phosphorylase Kinase Catalytic site Is any protein a target for the catalytic subunit of cAPK? No, only kinase enzymes will be acted upon. These target enzymes, therefore, must have some unique features. Phosphorylase kinase, for example, is composed of 4 different subunits (click 1). The delta subunit is calmodulin, a calcium binding protein, that regulates the activity (click 1). The gamma subunit has the catalytic site (click 1). The enzyme is not active. To activate phosphorylase kinase, the catalytic subunit transfers 2 PO4s from 2 ATPs to the alpha and beta subunits (click 1). Phosphorylation at these sites renders phosphorylase kinase active. Conversely, removing phosphate inactivates the kinase. Be alert, therefore, to phosphatases that may exert control over the function of phosphorylase kinases. Click 1 to go on.

5. Phosphorylase kinase cAMP dependent protein kinase Phosphorylase Adenylcyclase   R R   C C P P Crunch Time Now comes crunch time. What is the function of the target kinases. The answer is they phosphorylate enzymes that control a critical steps in a pathway. For example, phosphorylase or glycogen synthase are targets of a protein kinase. For phosphorylase, the kinase enzyme is phosphorylase kinase. The sequence of events is highlighted in the figure below (click 1). So now you can see how a hormone external to a cell can have a profound effect on glycogen degradation, glycogen synthesis, and other processes whose enzymes have activities controlled by the presence or absence of phosphate groups. Click 1 to test what you learned.

6. Test and Expand your Knowledge of cAMP Systems 1. Besides glucagon, can you name other hormones that elicit a cAMP response? Ans: epinephrine and norepinephrine 2. What name is given to a class of receptors that connect with adenylcyclase thru G proteins? Ans: Beta adrenergic 3. Based on the composition of phosphorylase kinase, what other factor may be expected to stimulate the activity of the enzyme and increase glycogen breakdown? Ans: Calcium ion. It binds to calmodulin one of the subunits of the kinase and activates the gamma (catalytic) subunit. 4. Supposing one hormone elicits 100 times it number of cAMP molecules. Then 2 cAMP molecules elicits 500 times their number of active phosphorylase kinases. Now suppose each phosphorylase kinase activates 1000 phosphorylase molecules. How many times has the single hormone been amplified? Ans: 25 million times. This means that one nanomole is having the same effect as 25 millimoles of hormone.