MCDB 310 1st Edition Lecture 14Outline of Last Lecture I. Examples of integral, trans-membrane Protein Functiona. Adhesionb. Fusion and Solute transferring (concentration and electrochemical gradients)II. Biochemistry in Cell SignalingOutline of Current Lecture I. Adhesion Receptors (continued from last lecture)II. Receptor Tyrosine KinasesIII. G-Protein Coupled ReceptorsIV. Non-steroidal hormone receptorsV. Nuclear receptorsVI. Guanylyl CyclasesVII. Principles of BioenergeticsCurrent LectureI. Adhesion Receptors: receptors for ligandsa. Ligands can be extracellular matrix components: weak interactions between protein and carbs induce changes outside cells (ligand/receptor interactions induce conformational changes)b. Extracellular conformational changes then extend to the protein domains inside the cells (leads to biochemical changes inside)c. The force (delta G) causes an ion to go through a channel is a function of the ration of the ion concentrations, the difference in membrane potential, and the number of open channelsi. Electrochemical gradient is often established by ACTIVE transportThese 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.ii. Then some solutes will take advantage of this gradient to move freely across a membraned. Voltage gated ion channels: Example-Neuronal Signalingi. Neurons move signals with 3 voltage gated channelsand 1 ligand gated channelii. Sodium (positive) enters the cell  depolarizing themembraneiii. Potassium channels meet their voltage threshold move potassium ions out  repolarizing themembraneiv. This repolarization induces another depolarization (3total voltage gated channels)v. At the junction of two cells, calcium bound vesiclesbind to the cell membrane and releases acetylcholine  diffuses to the next cell  received by aligand gated channelvi. This ligand gated channel causes depolarization cycle begins againvii. These channels change Vm:1. Locally2. In succession3. In one direction4. Via several channelse. Receptor Tyrosine Kinases (receptor enzymes, see insulin receptor figure as an example)i. A receptor + an enzyme in one proteinii. Ligand binding site on the extracellularface (example of ligand=insulin)iii. Single transmembrane domain perpolypeptide  conformational changeiv. Catalytic site on cytoplasmic face(inside) becomes activatedv. The catalytic activity isAUTOPHOSPHORYLATION (fairly rareoutside of these receptors)1. Three phosphoryl groups getadded to the catalytic domain  active2. No covalent changes, just weak interactions  conformational change swings the tyrosine into the active site of the enzymevi. There are many phosphorylation events inthis pathway (mostly on tyrosine) 1. Not the same activity in every cell,even with the same cell receptor2. Every time there is a conformationalchange (caused by phosphorylation),the enzymes can bind to somethingnew3. Eventually, one of these enzymes canget into the nucleus and cause aresponsevii. Phosphotyrosine is a ligandviii. The protein has a Phosphotyrosine BindingDomains (PTBs)1. Src Homology Domain 2 (SH2) is onesubclass of PTBa. High affinityb. Present on many signaling proteinsix. Regulation of this pathway: stop phosphorylation events1. This can happen at any step along the way because each enzyme has its own phosphorylation eventx. Low Molecular Weight G-Protein (Guanine nucleotide Binding) (unique to this pathway) called RAS1. These can bind GTP in active state and GDP in resting state2. There are two subclasses: low molecular weight and heterotrimeric3. These proteins have an active site to hydrolyze GTP4. Low molecular weight: tend to be smaller (one subunit) and soluble, named with 3 capital letters5. Heterotrimeric: 3 subunits (alpha, beta, and gamma), name starts with a G and then other letters that are indicative of function, associated with membranesxi. A kinase (ERK) moves into nucleus to phosphorylate the target protein, change the expression of protein (binds at major groove)xii. In this pathway there are NO secondary messengersf. G-Protein Coupled Receptors: using second messengersi. Plasma membrane receptor: one polypeptide with 7 trans-membrane domains (serpentine receptors)ii. Heterotrimeric G-Proteins that activate an enzymeiii. An enzyme in the membrane that generates one of more second messengersiv. Example: B-Adrenergic Receptor Mechanism1. Epinephrine binds its receptor  conformational change2. Activates the Stimulatory Heterotrimeric GTP binding protein3. GDP binds in its inactive state  it comes off  GTP binds to alpha subunit of dissociated G-Protein complex4. This activates adenylyl cyclase to make CYCLIC AMP (a second messenger)  protein kinase A is activated  phosphorylation5. A response: the response is AMPLIFIED (glucose is released into blood)v. Desensitization: shutting off signaling (stop and return the cell to previousstate to be stimulated again)1. Remove the ligand (epinephrine)2. Phosphorylate the alpha subunit of the G-Protein (inactivating it)3. Stops phosphorylation of second messengers4. Re-combine the G-protein5. Return the receptor to the cell surfaceg. Surface Hormone Receptors (also serpentine)i. Hormone (non-steroid) binds specific receptor on membraneii. Gq exchanges GDP for GTPiii. Gq then activates Phospholipase C with converts PIP2 to diacylglycerol and IP3 (both of which are second messengers)1. IP3 binds to a ligand gated channel  releases calcium into the cell  protein kinase C is activatediv. EITHER protein kinase C OR protein kinase A gets activated in one of thesesignaling pathways (never together in one cell)v. Summary: extracellular signals may cause different effects depending on the cell/tissue/second messengers releasedh. Phosphorylation as a regulatory mechanism (PKC and PKA)i. They regulated based on modules and rafts:1. Signaling happens through binding (docking) sites in groups of proteins (ie-Phosphotyrosine Binding Domain PTB)2. Modules: protein domain that is made up of a group of binding proteins (multivalent) (See table for examples)a. Build proteins with a bunch of units that each act as control points 3. Rafts (microdomains): regions of membranes enriched in certain porteins and lipids that can induce endo/exocytosisa. Different from a modulei. Binding Events and Scatchard Plotsi. Binding between two molecules can be difficult to analyze (transient and there