Biol 4610 1st Edition Lecture 13 Outline of Current Lecture 1. Molecular mechanisms of vesicular budding and fusion2. Forming of COPII vesicles3. Later stages of secretory pathway4. Endocytic pathway for transferrin receptorCurrent LectureSecretory Pathway à getting proteins from the ER to the Golgi, plasma membrane, secreted into extracellular space, lysosomesUsed for endocytosis*vesicle based transportAnterograde – forward transportRetrograde- backward transport14.2. Molecular Mechanisms of Vesicular Budding and FusionVesicles and formation of a protein coat – vesicle buds off from parents and fuses with a target membranev-SNAREs and t-SNAREs – vesicle SNARE and target snare- Recognizes where the vesicle is supposed to go to- V and t must match up with each otherFormation of COPII vesiclesMove from ER to early stages in GolgiThese 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.GTP binding proteins, such as Sar1, regulate coat assemblyReceptor in ER membrane, called Sec 12 (guanine exchange factor)Pull of GDP form a protein and allows GTP to bindDiacidic sorting signal (Asp –X-Glu) - A specific AA sequence that dictates what vesicle the protein is loaded into- The purple hook is the diacidic sorting signal - 14.3. Early Stages of the Secretory PathwayER to Golgi - Anterograde1. Cop III vesicles; Asp-X Glutamic acid2. Cargo proteins all have sorting signals- what directs the protein to go into a diff organelle; Diacidic sorting signal3. Target SNARES (receptor on target membrane)Golgi to ER – retrogradeThe KDEL (Lys-Asp-Glu-Leu ) sorting signal and COPI vesiclesRetrieve missorted soluble proteinsCisternal maturation – assembly line model1. Start at Cis-golgi compartment (close to ER)2. Enzymes change and cisternae move away form nucleus3. Modify carbohydrates on proteins4. End up at the trans-golgi network14.4. Later Stages of the Secretory Pathway- Trans-golgi network- Can have proteins that are secreted from the cells into plasma membrane (4)- Can become integral membrane proteins- Can form into a lysosomeClathrin vesicles – trans-golgi à endosome-endocytosis**Clathrin vesicles are all double coated**Lattice cage that surrounds the AP proteinsAP complexes (AP1, AP2, and AP3) –adaptor protein- Coat protein- Cargo binds to one of these AP proteins with specific sorting signalsDynamin- used to pinch off clathrin vesiclesSince two layers, clathrin can’t bud off on its own, must use Dynamin. Uses energy (GTP hydrolysis)Mannose 6-phosphate receptors-predominant use is to get proteins from trans-golgi into endosome-mannose 6 Phosphate tags proteins to go to endosome-mannose 6 Phosphate receptor binds to it- In Golgi- pH is 6.5; Endosome- pH is 5- So the lower pH lowers the affinity for the protein and releases it into the endosome1. Endocytic pathway for transferrin receptor- Iron is transported in the blood and bound to transferrin (carrier protein)- Ferrotransferrin à when it’s bound to iron- Apotransferrin à when it is NOT bound to iron- Ferrotransferrin binds to transferrin receptor on membrane- Sorting signal is in cytoplasmic domain, the binding of ferrotransferrin to receptor resultsin a comformational change to the protein which releases/exposes the sorting signal- YTRF ß Tyr-x-x-phi - The sorting signal can now be used by the AP2 proteins and polymerizes the second layer of clathrin (using Dyamin)- Reduced pH in endosome causes iron to be released by the transferrin molecule- Now it becomes Apotransferrin and recycles back up to plasma membrane- Finally it dissociates from the