BIOL 302 1st Edition Lecture 21Outline of Last Lecture I. Membrane transport pt 2Outline of Current Lecture I. ATP binding II. ABC transporters III. Single transporterIV. Single transduction Current LectureI. ATP-binding cassette (ABC) transportersA. ABC transporters: ATP binds to a specific domain, the ATP binding cassette (blue).B. One conformation when ATP is bound, another when ATP has been hydrolyzed and ADP is released. Hydrolysis of ATP is coupled to a conformational change.C. This type of pump does not get phosphorylated.II. ABC transportersThese notes represent a detailed interpretation of the professor’s lecture. GradeBuddy is best used as a supplement to your own notes, not as a substitute.III. ABC transportersIV. Ring cycle for dilating and constricting the nuclear poreV. Constricted and dilated transport channel of the nuclear poreVI. Signal transduction and G-protein coupled receptorsVII. Signal transductionA. An extracellular signal binds to its receptor. Example: a molecule of adrenalin, a hormone, binds to the β2-adrenergic receptor. B. The receptor is activated (or deactivated). Activation at the extracellular side needs to be transmitted into activation (or deactivation) at the intracellular side.C. Activation of the receptor initiates production of a secondary messenger inside the cell. This can be done via a pathway. Example: cAMP is produced or Ca2+ is released from its reservoirs. This amplifies the signal – one initial signaling molecule initiates release of a large amount of secondary messenger.D. The secondary messenger will reach and regulate several effectors in the cell thatdirectly alter the physiological response.E. Termination of the signal. After the cell has responded to a signal, the signaling process must be terminated or the cell loses responsiveness to new signals.VIII. Secondary messengersIX. G-protein coupled receptorsA. There are over 800 GPCRs in the human genome and they respond to a broad spectrum of chemical entities ranging fromphotons, protons and calcium ions, small organic molecules (including odorants and neurotransmitters), to peptides and glycoproteins.B. The classical role of a GPCR is to detect the presence of an extracellular agonist, transmit the information across the plasma membrane, and activate a cytoplasmic heterotrimeric G protein, leading to modulation of downstream effector proteins.X. Rhodopsin and β2-adrenergic receptorXI. A trimeric G-protein in GDP fromA. G-proteins are heterotrimeric and consist of α, β and γ subunits. Gα has an intrinsic GTPase activity and can exist in GDP (inactive) and GTP (active) bound states.B. In the GTP state, Gα dissociates from the Gβγ dimer. XII. G-protein coupled receptorsA. The extracellular signal (an agonist) is transmitted into an intracellular signal. B. G protein is activated by exchange of GDP to GTP, which is mediated by the GPCR-agonist complex. After the G-protein is complexed with GTP, it dissociates into Gα and Gβγ subunits. Gα can activate Adenylate cyclase for signaling, which makes cAMP. However, activated Gα hydrolyses GTP slowly and thus deactivates itself.C. Gs – stimulatory G protein – activates Adenyl cyclase - binds to GPCR-agonist complexD. Gi – inhibitory G protein – inhibits Adenyl cyclase - binds to GPCR-inhibitor complexXIII. Activation of protein kinase A by a G-protein pathwayA. Adenylate cyclase is activated by the α subunit of the G protein in the GTP form.B. Adenylate cyclase is a membrane protein that synthesizes cAMP (cyclic AMP) from ATP.C. cAMP is a secondary messenger that activates protein kinase A and other targetsXIV. Resetting the G proteinA. On hydrolysis of the bound GTP by the intrinsic GTPase activity of Gα (which slowly happens by itself), Gα reassociates with the βγ dimer to form the heterotrimeric G protein, thereby terminating the activation of adenylate cyclase.XV. Signal termination – the Arrestin pathwayA. G-protein Receptor Kinase (GRK) phosphorylates the GPCR on its cytoplasmic C-terminal tail. This increases its affinity for a protein called beta-arrestin (red), thatbinds to the receptor and prevents it from associating with a trimeric G-protein. Beta-arrestin can lead to desenzitation or target the receptor for endocytosis, butit also acts as signaling molecule in its own right.XVI. G-protein coupled receptorsA. The complex signaling and regulatory behavior of the β2AR. B. PKA=protein kinase A, PKC=protein kinase C, PDE=phosphodiesterase, cAMP=cyclic adenosin monophosphate, Gs=stimulative regulative G protein, Gi=inhibitory regulative G protein, GRK=G-protein-coupled receptor kinase, ERK=extracellular signal-regulated kinase.XVII. G-protein coupled receptorsA. Many GPCRs have more complex signaling repertoires than the classical pathway.For example, the β2AR couples to both Gαs (stimulatory G protein) and Gαi (inhibitory G protein) and can also signal through MAP kinase pathways in a G-protein-independent manner via arrestin. Similarly, the process of GPCR desensitization involves multiple pathways, including receptor phosphorylation events, arrestin-mediated internalization into endosomes, receptor recycling, andlysosomal degradation.XVIII. Lipid rafts as membrane-organizing principle (Kai Simons)A. Lipid rafts: Cholesterol and SphingolipidsB. These lipid rafts form a ‘compartment’ within the membrane as they phase-separate fromPhospholipids1. Cholesterol induces lateral phase separation into domains in the membrane containing glycerophospholipids and sphingolipids. GPL stands for glycerophospholipid. The grey–yellow part of the membrane at the bottom represents a raft. Below: formation of rafts in a lipid bilayer upon the additionof cholesterol. Lα stands for a lamellar phase and lo is the ordered lipid domain.XIX. G-protein coupled receptorsA. First crystal structures of β2AR in the inactive states. A) The β2AR–Fab complex. B) The β2AR–T4L fusion protein.XX. G-protein coupled receptorsA. In the absence of a ligand, the G protein coupling interface of the receptor exists in an ensemble of predominantly low-energy conformations. Rare active-state conformations are responsible for basal activity. Agonist binding increases the dynamic properties of the β2AR, increasing the probably of active state conformations. Only G protein binding can fully stabilize the active state.XXI. G-protein coupled receptorsA. A comparison of the carazolol-bound, inactive-state structure