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What goes on in the ER?
- - Biosynthesis of core oligo. for N-linked glycosylation
- - attachment of core oligosaccharide to asparagine residues
What goes on in the CGN?
- - attachment of N-acetylgalactosamine to serine or threonine
- - first step of phosphorylation of lysosomal proteins
What goes on in the Cis region?
- - removal of mannose
- - second step of phosphorylation of lysosomal proteins
What goes on in the medial region?
- removal of mannose
- attachment of N-acetylglucosamine
What goes on in the trans region?
- addition of galactose
- addition of sialic acid
What goes on in the TGN?
- addition of sialic accid
- atachment of sulfate to tyrosine
Two pathways of protein secretion?
- constitutive: newly synthesized molecules are not concentrated; there is no post-Golgi storage pool; and te transport vesicle has a short transient time from Golgi to surface
- regulated: the secretory products are condensed in electron-dense core forms; the dense core vesicles accumulate in the cytoplasm becuase they cna remain there for an extended period of time; fusion requires an intracellular messenger
In constitutive secretion (aka: __), protiens do not __. There are no __. There have to be __, but no __ nor __.
- continuous secretion
- get concentrated
- condensing vacuoles like digestive cells
- external signal
Nonpolarized vs. Polarized
- nonpolarized: unregulated/ the type of cell determines what is produced; non-specific binding or receptors involved; uses constitutive-pathway
- Polarized: some cells are regulated; others are not; it will only fuse in very specific places; contain a different sorting sequence if destined to apical surface then those of hte lateral or basal layers (special sorting signals used); selectively directed
From the trans face, three pathways can take place. What are they?
Synthesis of cholesterol
- - the two membrane-bound enzymes studied are HMG reductase and squalene synthesase.
- - Enzymes embedded in the SER have their active sites facing the cytosol so that the product ends up in the cytosol
Conversion of cholesterol to steroid hormones
- - intermediates get shuttled back and forth between the organelles to complete appropriate transformations
- - The two ways that it can begin is with cholesterol going into the mitochondrion of the adrenal cortex or acetate going into the adrenal cortex and being converted to cholesterol by the smooth ER, then allowing entry into the mitochondrion
- - All cells of the adrenal cortex carry out the pathway to progesterone
- - The formation of cortisol occurs in the inner cortex, while corticosterone and aldosterone are made in the glomerulosa
Process of centrifugation
- - break cells open and homogenize them
- - place in isotonic sucrose solution and crush with pestle; a blender may also be used ot break them open; this is going to homogenize the tissue fragment
- - take the homogenate and centrifuge it at a slow speed for a couple of minutes
- - remove the supernatant, leaving just the pellet of heavy particles
- - take the supernatant with the particles, centrifuge at a higher speed, getting a new pellet
- - take the supernatant off, leaving just the pellet
- - centrifuge the supernatant, getting more tiny particles
- - layer one on top of the other
- o sedimentation will depend on their density in relation to the gradient
Why is the cisternal maturation model the consensus?
- - It envisions a highly dynamic Golgi complex in which the major eleemnts of the organelle, the cisternae, are continually being formed at the cis face and dispersed at the trans face. The existence of the Golgi depends on continual influx of transport carriers from the ER and ERGIC
- - Certain materials that are produced in the ER and travel through the Golgi are shown to remain in cisternae and not appear in vesicles
- - The new idea that vesicles can move in a retrograde direction from trans donor to cis acceptor
- - Composition of the Golgi changes over time
- 1) Sar1 recruited to the membrane in GDP bound form and is induced to exchange GDP for GTP, causing a conformational change that causes its N-terminal alpha helix to insert itself intot he cytosolic leaflet of the ERà bends lipid bilayer
- 2) Recruitment of two additional polypeptides causes, due to the curved shape, additional pressure on the membrane surface to help it further bend into a curved bud
- 3) The remaining subunits of the COPII coat bind to the membrane to form the scaffold fo the protein coat
- 4) Once assembled, it buds to form the COPII-coated vesicle
- 5) Protein coat disassembled before fusion to the membrane and release into the cytosol
- a. Disassembly triggered by hydrolysis of the bound GTP to produce a Sar1-GDP subunit
COPI transport functions in what
(retrograde transport of proteins [Golgià ER or trans faceàCis face])
Molecules with an ER retrieval signal (the KDEL: lys-asp-glu-leu) will be captured and returned to the ER in COPI-coated vesicles