Chem 454: Regulatory Mechanisms in BiochemistryUniversity of Wisconsin-Eau Claire

Signal Transduction:Information Metabolism

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Information MetabolismHow cells receive, process and respond to information from the environment.

A large number of proteins are involved in information processing

Signal-transduction cascadesMolecular circuits that detect, amplify, integrate external signals.

Produce changes in enzyme activity, gene expression or ion channel activity.

Introduction

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Signal transduction depends on molecular circuits

Overview

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Transferring information from the cell’s exterior to it’s interior involves

Primary messenger

Cell surface receptors

Formation of a receptor-ligand complex

Production of a second messenger within the cell

This is transduction

Overview

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Examples of second messengers

Overview

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Protein phosphorylation can occur in response to the second messenger.

Overview

Tyr

Tyr

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Signal termination

Without signal termination, cells lose their responsiveness to the signal

Some cancers are associated with the inability to properly terminates signals

Overview

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Essentially every biochemical process in a cell is a component of a signal-transduction pathway or can be affected by one.

Overview

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The seven-transmembrane-helix receptors (7TM) contain a bundle of 7 α-helices, which span the cell membrane

1. Seven-Transmembrane-Helix Receptors

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The seven-transmembrane-helix receptors (7TM) respond to a wide range of primary messengers

1. Seven-Transmembrane-Helix Receptors

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For β-adrenergic receptors, ligand binding to 7TM receptors leads to the activation ofG-proteins

1.1. Ligand Binding to 7TM Receptors

HO

HO

N CH3

HHO H

epinephrine (adrenaline)

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G-Proteins cycle between GDP- and GTP-bound forms

1.2. G-Proteins

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G-Proteins cycle between GDP- and GTP-bound forms

1.2. G-Proteins

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G-Proteins cycle between GDP- and GTP-bound forms

1.2. G-Proteins

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There are many types of G-proteins

1.2. G-Proteins

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Activated G-proteins transmit signals by binding to other proteins.

For example, with the β-adrenergic receptors, the activated G-protein (α-subunit) binds to, and activates, the enzyme adenylate cyclase.Adenylate cyclase catalyzes the synthesis of cyclic-AMP from ATP

Cyclic-AMP is a second messenger

1.3. Activated G Proteins

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G-proteins spontaneously reset themselves through GTP hydrolysis

1.4. Resetting G Proteins

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For β-adrenergic receptors, prolonged exposure to epinephrine leads to desensitization of the receptor by β-Arrestin

1.4. Resetting G Proteins

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Cyclic-AMP stimulates the phosphorylation of many target proteins by Protein Kinase A

Processes that are stimulated includeEnhanced degradation of storage fuels

Increased secretion of acid by gastric mucosa

Diminished aggregation of blood platelets

Opening of chloride channels

1.5. Cyclic-AMP

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Phosphorylation of enzymes can turn them off or on.

1.5. Cyclic-AMP

Tyr

Tyr

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Activation of the enzyme phospholipase C by a G-protein produces two other second messengers

Phospholipase C cleaves phosphotidyl inositol 4,5 bisphosphate to produce

inositol 1,4,5-trisphosphatediacylglycerol

2. Phopholipase C

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2. Phopholipase C

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Inositol 1,4,5-trisphosphate opens channels to relases calcium ions from intracellular stores

The Ca++ is released by the stores in the endoplasmic reticulum or the sarcoplasmic reticulum (smooth muscle)

The Ca++ triggers smooth muscle contraction, glycogen breakdown, and vesicle release

2.1. Inositol 1,4,5-Trisphosphate

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Diacylglycerol activates protein kinase C

2.2. Diacylglycerol

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Calcium ion is an ubiquitous cytosolic messenger

The cytosolic centraction of Ca++ ion can by changed very rapidly.

The cytosolic concentration (100 nM) is very low compared to the extracellular concentration (5 mM)

Ca++ can bind tightly to proteins

3. Calcium Ion

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Calcium (second messenger) activates the regulatory protein calmodulin

Calmodulin contains a couple of EF-hand calcium ion binding motifs

3.2. Calmodulin

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Binding calcium converts calmodulin to an active form

3.2. Calmodulin

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The active form of calmodulin binds to and alters the activity of a wide range of cellular proteins

3.2. Calmodulin

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Calmodulin activates the calcium-dedpendent protein kinase (CaM kinase I) by binding to a C-terminal α-helilx

3.2. Calmodulin

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Some receptors dimerize in response to ligand binding

The dimerization brings together protein kinases that are associated with the intracellular domains of the receptors.

The protein kinases are then activated by cross-phosphorylization

4. Cross Phosphorylation

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Example: Human growth hormoneDimerizes the human growth hormone receptorWhich in turn, activates Janus kinase 2 (JAK2)

4. Cross-phosporylation

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Example: Human growth hormoneDimerizes the human growth hormone receptorWhich in turn, activates Janus kinase 2 (JAK2)

4. Cross-phosporylation

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Example: Human growth hormoneDimerizes the human growth hormone receptorWhich in turn, activates Janus kinase 2 (JAK2)

4. Cross-phosporylation

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The JAK 2 kinase has a modular domain structure:

4. Cross-phosporylation

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The SH2 domain is a regulatory domain that binds to phosphotyrosine-containing polypeptides:

4. Cross-phosporylation

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Learn more about the SH2 domain:The Structural Insights module on SH2 domains describes some of the determinants of SH2 specificity and the ways in which SH2-phosphotyrosine binding can affect protein function. Given that the Src kinase SH2 domain bind Src Phophotyrosine 527, what effect do yo think mutation of Glu 529 to Asn would have on th protein kinase activity of Src? Suppose you now obtained a second mutation within Src that reversed the effect of the first. Can you predict what that second mutation might be?

Problem 15.16

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When activated, JAK2 phosphorylates other proteins

The growth hormone receptorSTAT5, which regulataes gene expression

4. Cross-phosporylation

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Activation of growth hormone receptor by JAK2

4. Cross-phosporylation

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Activation of the STAT5 gene regulator protein by JAK2

4. Cross-phosporylation

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Some receptors contain tyrosine kinase domains within their covalent structures

The epidermal growth factor receptor is an example

4.1. Tyrosine Kinase Receptors

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The EGF signal transduction pathway includes a number of players.

4.1. Tyrosine Kinase Receptors

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The Grb-2 adaptor protein contains an SH2 and two SH3 domains

4.1. Tyrosine Kinase Receptors

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The Grb-2 adaptor protein contains an SH2 and two SH3 domains

4.1. Tyrosine Kinase Receptors

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The Sos proteins facilitates the exchange of GTP for GDP in the Ras Protein

4.1. Tyrosine Kinase Receptors

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The Ras protein is a member of a superfamily of monomeric G-proteins

4.2. Ras Protein

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Defects in signaling pathways can lead to cancer

Cancer is associated with uncontrolled cell growth.Cancer is strongly associated with signal transduction proteins

5. Signaling Defects and Disease

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Example Rous Sarcoma Virus codes for a protein, v-Src, which is homologous to the cellular, c-Src.

Rouse Sarcoma Virus produces sarcomas in chickens

The v-Src protein induces cell transformation, whereas the c-Src normally does not.

5. Signaling Defects and Disease

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The c-Src contains an SH2, an SH3 and a tyrosine kinase domain.

5. Signaling Defects and Disease

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The inactive c-Src can be activated in three different ways

5. Signaling Defects and Disease

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Protein kinase inhibitors may be effective anticancer drugs

90% of patients with chronic mylogenous leukemia (CML) have a specific chromosomal defect that results in a mutant protein kinase (bcr-abl) that is expressed in levels higher than they should be.

Research into protein kinase inhibitors is showing probmise

5.1. Protein Kinase Inhibitors

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Protein kinase inhibitors may be effective anticancer drugs

5.1. Protein Kinase Inhibitors