Cell Biology Exam 2 Chapter 14 Steps in secretory pathway o Import into ER o Folding and glycosylation in ER o Vesicular transport from ER to Golgi o Cisternal progression through golgi glycosylation other mods here o Vesicular sorting at trans golgi network o Vesicular trafficking to final destination Plasma membrane Lysosome Endosome Vesicular Transport from ER and Golgi Transport o Membranes fuse and segments are exchanged o Vesicle cytoplasmic conditions and contents vary but the resident enzymes in the ER always remain the same o Types for both membrane bound and soluble proteins Anterograde from ER to cis golgi network COP2 coat grabs hold of the membrane of ER to create a vesicle for transport filled with cargo to golgi Retrograde cis golgi back to ER COP1 coat grabs hold of the membrane of golgi to create a vesicle for transport filled with cargo to ER o Retention signals KDEL keeps resident proteins in the ER Lys Asp Glu Leu Coo On c terminal end Binds to KDEL receptors on golgi membrane in high ion concentrations The protein is then internalized in vesicles in retrograde transport and is moved back to the ER The receptor releases the protein back into the ER because of the low ion concentrations in the ER and it is recycled back to the golgi o ERGIC Compartment another name for cis golgi Vesicle Budding Process o Coatomers bind to the membrane coat proteins o GTP binding protein o V Snares help to bind to the target membrane Matches T snares on the membrane o Cargo Membrane cargo protein embedded in bilayer Soluble cargo protein grabbed by membrane bound cargo receptors o GTPase ARF with COP2 Sar1 with COP1 Caltherins also use this o Sar1 which is a soluble protein in cytosol binds to the SEC 12 receptor on the ER membrane Sar1 carries GDP o SEC 12 is a GTP GDP exchange factor When SAR1 binds then the GTP is thrown off and the GTP is added When GTP adds it undergoes a conformational change and the tail structure on Sar 1 is embedded in the cytosolic membrane o SEC23 SEC24 coat proteins bind to their receptor Sar1 when the tail is embedded Process o The shape of the proteins bulges out the section of the membrane creating a vesicle o After it is pinched off Sar1 hydrolyzes its GTP to GDP The tail pulls out and the green coat proteins begin to fall off Result is a naked vesicle Directing proteins to vesicles o Luminal sorting signals soluble proteins in the lumen carry KDEL on the C terminal end o Cytosolic sorting signals on the C terminal end of the proteins embedded in the KKXX in the lumen membrane KKXX in the cytosol o KKXX flag for the proteins on the C terminal end Fusion of vesicles with their target membranes o Coat is already thrown off o GTPase remains Rab GTP o VAMP v snare vesicle associated membrane protein on vesicle o T snares on target membranes Syntaxin SNAP 25 o Rab GTP binds to Rab effector Hydrolyzes for energy to pull the membrane very close In order to fully pull the t and v snares together they coil together final pull onto the membrane coiled coil structure When the membranes touch they fuse and the soluble contents are released into the golgi and the membrane proteins become a part of the golgi o After contents are released the coiled coil structure remains on the membrane of the golgi NSF and alpha snap hydrolyze ATP and unravel t snare and v snare so that they can be reused o Mechanism acts in both anterograde and retrograde transport Caltharin Coats o They take proteins from the trans golgi to the plasma membrane o Has a triskelion structure Heavy chains Light chains Globular head embedded in the structure Dynamin GTPase that uses the energy from the hydrolysis of GTP to wrap around the thin part of the forming vesicle Continues to use GTP until fusion and a complete vesicle is formed Cisternae stacks of the golgi classification of the stacks is dependent on the resident enzymes that they contain Cisternal Progression Through Golgi o Cis golgi network ERGIC o Cis golgi o Medial golgi o Trans golgi o Trans golgi network Sialic Acid o When you take a neuraminic acid and you either substitute N or O o Nine carbon backbone that usually occurs at the end of sugar chains o Example N acetyl neuraminic acid o Important because the influenza virus uses neuraminidase enzymes N1 N2 etc o Receptors on the membrane of cells recognize sialic acid Cells recognize the neuraminidase enzymes on the virus and the virus enters the cell via endocytosis Glycosylation adds diversity to proteins o Roles o N Linked Folding addition of oligosaccharide maintains proper folding Resistance to proteases stabilizes and increases the half lives Protein Protein interactions Protein ECM interactions provides strength Targeting act as signaling molecules in the cytosol Enzyme Regulation antibodies use glycosylation to allow them to stick to pathogens and activate enzymes Sugar is joined onto the nitrogen on the asparagine In order to be N liked it must have ASP X THR SER X If there is a proline in the X group the oligosaccharide will not join Entire oligosaccharide with dolichol was added en masse oligosaccharyl transferase Maturation Cis cisterna removal of 3 mannose Medial cisterna add an GlcNAc with UDP by GlcNAc transferase 2 mannose removed by mannosidase two GlcNAc added again by UDP fructose is relocated by GDP Trans golgi addition of 3 galactose with UDP addition of 3 NANA with CMP Glycan sugar group called proteoglycan when bound to a protein o O linked Two classes of mature N linked Oligosaccharides High Mannose terminal end is filled with mannose Complex goes through a high mannose intermediate negatively charged Endoglycosidase H enzyme that chops up the mannose when exposed to it complex n linked are resistant because of the other sugar groups attached to the mannose that shield it from the enzyme Sugar is attached to the OH group on the threonine or the serine Requires X SER THR X If there is a proline in the X group the oligosaccharide would love to join Sugars are added one by one not en masse Added by glycosyl transferase OH enters and grabs the H off of the hydroxyl group Water exits and the protein binds to the left over O with a negative charge Very important in the extracellular matrix Mucins Lungs nose stomach intestines where you are exposed to the environment is lined with mucous for protection by filtering out the debris and for defense against dehydration mucins o linked sugar o A proteoglycan that is heavily branched to trap water Collagen o linked