MIDTERMS AAAAAAAAAAAAAAAA Flashcards
1
Q
What takes up most of a cells mass except for water
A
protein
2
Q
What percent of genes are shared between human cells
A
30-60
3
Q
Do you need a primer for transcription
A
no you idiot
4
Q
What is the abortive transcription
A
When 10 nucleotides are formed, sigma factor is released. This can happen in eukaryotes but with general transcription factors instead of sigma factor
5
Q
What is a hairpin structure
A
A looping of mRNA that happens in transcription which causes the RNA polymerase to leave the DNA strand
6
Q
What are gene regulatory proteins or transcription factors? What do they bind to, specifically?
A
They regulate gene expression by binding to regulatory sequences of DNA called cis factors.
7
Q
What are two types of gene regulatory proteins/transcription factors
A
Activators and repressors
8
Q
What is an operon
A
multiple genes transcribed into one RNA molecule
9
Q
What is the Trp operon, how does the trp repressor work
A
5 genes encode enzymes for tryptophan synthesis, transcription regulated by a single operator. the Trp repressor binds to the operator sequence to prevent RNA polymerase from binding. the trp repressor binds 2 molecules of tryptophan to bind to the helix-turn helix binding motif in the major groove of DNA
10
Q
What is the most common DNA binding motif
A
the binding of a gene regulatory protein in the major groove of DNA (such as with trp operon)
11
Q
Explain the lac operon in E coli. What does Ecoli prefer
A
Ecoli uses lactose when there is no glucose. The operon is turned on when using lactose. However, E coli prefers glucose
12
Q
What activates the lac operon, under what condition
A
Catablite Activator Protein (CAP) which increases when glucose decreases. This increases cAMP which changes CAP conformation, and allows it to bind to DNA. It binds to promoters to help yoink RNA polymerase and get RNA poly. to bind.
13
Q
What represses the lac operon and under what conditions
A
Lac repressor protein shuts off the operon when there is no lactose. This is because allolactose increases as lactose increases, releasing lac repressor from the operator (conformational change).
14
Q
What does DNA sequencing, DNA microarrays, 2D gel electrophoresis show you
A
Respectively, info about DNA, RNA, and proteome
15
Q
What is negative and positive regulation in prokaryotic transcriptional regulation
A
Repressor preventing activity, and activator promoting activity
16
Q
What are the two ways ligands can control gene-regulating proteins
A
Ligands can yoink regulatory protein from the DNA or yeet the regulatory protien onto it
17
Q
Where can regulatory elements be found
A
Upstream or downstream of the promoter. Downstream and within genes is common for eukaryotes.
18
Q
What is kissing :3
A
I’m a visual learner btw (jk its when distant regulatory elements like NRTC loops back allowing NRTC to interact with RNA polymerase at a distance. can happen in both eukaryotes and prokaryotes)
19
Q
Explain how the lambda repressor and cro protein work
A
The lambda repressor binds to the cro side of the operator, which activates its own synthesis, and most bacteriophage DNA is not transcribed. The cro protein binds to the other side and activates its own synthesis as well. It’s also a repressor. They both essentially prevent transcription from one side
20
Q
Explain the prophage state of lambda DNA after it enters the host
A
Lambda DNA is put into hist genome after lambda repressor binds to the operator. The cell divides and this leads to lambda DNA dividing with host DNA (it gets a free ride)
21
Q
Explain how a bacteria may switch from prophage to lytic pathway
A
When the host reacts to DNA damage
22
Q
Explain the lytic pathway after lambda DNA enters host
A
Cro blocks lambda repressor, and bacterial DNA is extensively transcribed, so theres a shit ton of viral proteins needed for formation of new viruses. Then lambda DNA gets replicated and gets put into viruses. Cell lysis, and the viruses leaves >:) muahahahahah
23
Q
What promotes prophage state in bacteriaphage infected bacteria
A
If conditions are good for growth and cell division, then lambda repressor turns off cro, maintaining prophage state.
24
Q
What is a transcriptional circuit (simply?)
A
When A protein is coded for by a certain piece of DNA, and this A protein then goes back and either activates or represses the operator.
25
Explain the flip flop device
One DNA codes for A which stops B synthesis. The B DNA stops A synthesis. Its also known as indirect positive feedback loop
26
Explain feedforward loop
Protein A activates B and Z synthesis. Protein B activates Z synthesis. Z is super duper ultra mega activated >:)
27
Explain cell memory in the context feed forward loops
A cell has a gene, and initial transitent signal turns the gene on. Now you make the protein. The protein facilitates its own creation (positive feedback) so its sons and daughters and children all have the protein and have the gene activated
28
How can feedforward loops be used to measure duration of a signal
Both A and B are needed to activate Z, so the sensitivity of Z is decreased. output of Z is only achieved with prolonged input.
29
What is synthetic biology
Constructing artificial regulatory circuits and examining behavior in cells
30
What is a repressilator, what happens with the protein synthesized IRL
A is expressed, B repressed, C expressed. C then represses A which expresses B, repressing C. It's like a bunch of people who really want to fuck with everyone else. IRL the protein amount oscillates up and down but overall goes up due to there being more overall bacteria
31
What is transcription attenuation
Present in both eukaryotes and prokaryotes. RNA gets a structure that fucks with RNA polymerase and Regulatory proteins can bind to RNA and interfere with attenuation. Prokaryotes, plants, and some fungi use riboswitches to regulate gene expression.
32
What is a riboswitch
(Someone who isn't a ribotop or a ribobottom /j) short RNA sequences that change conformation when bound by a small molecule. some do transcription attenuation, some do stuff other than transcription attenuation. an example is when high purine, purine binds to the riboswitch, changing it conformationally, and then RNA polymerase is pried off.
33
What do general transcription factors do, name the 5 needed by RNA polymerase 2
help position eukaryotic RNA polymerases at eykaryotic promoters. The 5 are TFIID, TFIIB, TFIIF, TFIIE, TFIIH.
34
What does RNA polymerase II code for
protein coding genes
35
What does the mediator do
correctly positions TFIIH, influences general transcription proteins
36
What do elongation factors do
Prevent DNA polymerase from dissociating
37
What are coactivators and corepressors
Shit that assemble on gene regulatory proteins but don't directly bind to DNA
38
What are the two parts of a eukaryotic activator protein, how are they positioned on the DNA
DNA binding domain (DBD) recognizes specific DNA sequence. Activation domain (AD) actually accelerates transcription. DBD is the foot part and AD is the doorknob part
39
How do activator proteins facilitate transcription
Attracting, positioning, modifying general transcription factors, mediators, RNA polymerase II. They either directly act on components like prokaryotic activators (e.g. bind somewhere on DNA and succc the thing it wants close) or by indirectly modifying chromatin structure
40
How many base pairs around neocleosomes, how many times around octamer core, how long is linker DNA (just like get the general range)
Nucleosomes are 147 bp, wound 1 2/3 around octamer, linker DNA 10-80 bp
41
What are the 4 major ways activator proteins activate chromatin
Remodelling nucleosomes use chromatin remodelling complex, using ATP to pull DNA along nucleosome. Histone removal, histone replacement (need histone chaperones), specific patterns of histone modification
42
What is the histone code
Acetylation, methylation, phosphorylation on histone tails. Modifications are dynamic and determined by DNA. code readers provide meaning
43
What termini are histone tails
Histone tails are usually N-termini, C termini can stick out too but its rare
44
How does eukaryotic repression happen
Competitie DNA binding, masking activation (the thing where the doorknobs are fighting), direct interaction with general transcription factors, recruitment of chromatin remodelling complexes, deacetylation by recruitinng deacetylases, methyl transferase recruiting.
45
How do reader writer proteins stabilize chromatin
A reader protein recruits writer which starts domino effect of reader-writers complex. DNA methylase enzyme is attracted by the reader which methylates nearby cytosines, stabilizing chromatin.
46
Methylation of chromatin and blocking of gene expression as a result is an example of what
epigenetic inheritance
47
How does the human interferon gene have regulation
Activator attracts histone acetyltransferase, acetylates H4K8, H4K9, activator protein attracts histone kinase that phosphorylates H4S10, and serine modification signals the acetyltransferase to acetylate H3K14. Remember that there are 2 tails for H3 because there are 2 H3s
48
What is RNA capping
Addition of modified guanine to the 5' end of pre-mRNA, distinguishing it from other types of RNA. Done by cap binding complex that prevents 5'exonuclease from cutting phosphodiester bonds (it cannot cut the weird guanine)
49
What is splicing done by
The spliceosome
50
What marks proper splicing
EJCs
51
How is the sex lethal gene regulated
The splice site is blocked, which makes a shit ton more spliced. Like the stuff between two introns are spliced out too
52
How is the transformer gene regulated
Sex lethal makes Sxl protein which blocks the splice site and makes an intron longer
53
How is the doublesex gene regulated
The transformer gene creates a protein that activates a splice site, which prevents the exon from being spliced (best to double check with slides for the drosophila related genes because the diagrams help a lot)
54
What makes a male drosophila
Nonfunctional Sxl, nonfunctional Tra, inactive 3' splice for doublesex
55
How is eukaryotic transcription terminated
CstF (cleavage stimulating factor) and CPSF (cleavage and polyadenylation specificity factor) are on RNA polymerase tail and transfer onto 3' processing sequence. RNA get cleaved, poly A polymerase adds a ton of not genome encoded As.
56
What binds RNA processing proteins before they go to RNA
C-terminal domain of RNA polymerase
56
What causes the RNA processing proteins to be yeeted on
Phosphorylation of RNA polymerase
57
What are markers of mature RNA you need to move it out of the nucleus
Cap binding complexes, exon-junction complexes, poly A binding proteins.
58
What can you not have on proteins for export
snRNPs must be lost for export
59
What happens to messed up RNAs that aren't fit for export? how much RNA leaves the nucleus
1/20 leaves the nucleus. messed up RNA will be degraded in the nucleus by the exosome.
60
How do eukaryotes make sure the mRNA is good quality once it is exported out of the nucleus?
5' cap bound by elF4E, poly A binding protein bound by elF4G. These 2 recruit small ribosomal complexes to start translation. This makes sure both ends of the RNA are intact
61
What is nonsense mediated mRNA decay? What should happen if the splicing is normal?
Normal conditions will have EJCs displaced by moving ribosome. Stop codon is on the last exon. Abnormal splicing will cause a premature stop codon which leads to EJCs still on RNA. UPF degrades fucky wucky mRNA
62
Where is nonsense mediated mRNA decay important
In the immune system where a lot of DNA rearrangement happens to make antibodies.
63
How do prokaryotes control for mRNA quality
Ribosomes get stuck on broken and incomplete RNAs, and a special tmRNA (好礼貌的rna草) gets yoinked to the A side, and puts alanine onto the chain. The tmRNA acts as the template for the next tRNAs, making an 11 amino acid tag, and proteases will be attracted to shit with the tag and eat it.
64
How does the deadenylase act as a timer of mRNA lifetime
It is a 3-5 exonuclease. Once the polyA tail is too short, then the 5' end can be dropped, or 3-5 degradation continues. Or both.
65
How can mRNA be stabilized in the cytoplasm
cytoplasmic poly-A elongation. some proteins can fuck with this tho
66
What does transferrin receptor do
yeets iron into the cell
67
How is transferrin regulated by mRNA
mRNA is stabilized by cytosolic aconitase when there is low iron in the cell, allowing the transferrin receptor to be made. when there is high iron, the 3'UTR endonucleolytic side is exposed because the iron sticks to mRNA.
68
How are miRNAs made
synthesized by RNA polymerase II and base pairs with itself. the parts not base paired are cleaved in the nucleus, then the dicer removes single parts in the cytosol. Initially, it has a 5' cap and poly-A tail. it then yoinks a argonaute to form a RISC (RNA-induced silencing complex)
69
What is argonaute
A protein that helps miRNAs find targets. The target is either eaten, or it blocks translation, which takes a while, and then usually degrades the RNA.
70
What is RNAi
Double stranded RNA that blocks other RNA in a sequence specific manner, like miRNAs
71
What are siRNAs
small interfereing RNA. double stranded, can work like miRNA or interact with newly transcribed RNA, recruit chromatin modifying enzymes to compact DNA and then no more translation
72
How does crispr-cas work
Bits of viral DNA get into CRISPR locus, then it makes pre-crRNA from the DNA sequences that are acquired from viral DNA. The pre-crRNA is processed and added onto Cas protein (which acts like argonaute). Then the cas protein and crRNA go to destroy complementary viral DNA
73
What does shine-dalgarno sequence do
a sequence upstream of aug start codon (5'UTR). this correctly positions aug in ribosome and provides translational control
74
What are the four ways that the shine dalgarno sequence can be regulated
1. RNA binnding protein blocks access to it, so no protein made. 2. a stem-loop RNA structure blocks access to the shine-dalgarno sequence 3. riboswitch causes structural rearrangement of DNA, blocking Shine-Dalgarno sequence. 4. antisense RNA made somewhere else base pairs with mRNA, blocking SD. this antisense RNA is not miRNA, siRNA, or crRNA
75
How is translational regulation regulated in eukaryotes
Translational repressors can bind near the initiator AUG and inhibit translation. Like how ferritin binds iron, which yoinks the aconitase off of the RNA, allowing ferritin to be made. When iron is low, there is no iron to yoink it off, meaning no ferritin is made. Repressor proteins can also interfere with 5'cap and 3' poly A tail, which are both needed for efficient translation.
76
How can you modify eukaryotic initiation factors (elFs) to regulate translation
Phosphorylate them. elF2 forms a complex with GTP, recruits methionine tRNA to small ribosomal subunit. When the mthionine carrying tRNA finds AUG, ATP is hydrolysed to ADP. This changes elF2 shape, and yeets it out with a GDP attached. GDP attached elF2 is inactive and needs elF2B to activate it. Phosphorylating elF2 makes elF2 stuck to elF2B, and now elF2B cannot go do its job :( now translation is very slow. not all mRNAs are equally affected by this
77
Examples of post translational regulation
proteins must fold properly, so you can have covalently modified proteins (sugar, phosphate), proteins interacting with other proteins and small molecules (cofactors). some protein folding begins as soon as they emerge from ribosomes and some after synthesis
78
What is another name for chaperone proteins
heat-shock proteins because a shit ton is made when its hot. they are also there when its normally hot
79
How do Hsp70 and 60 fold proteins
they interact with exposed hydrophobic parts of proteins, which is not normal, obviously, and use ATP to fold them in one of 2 chambers. theres a lil cute lid to keep stuff in place and prevent shit from getting mixed up
80
What happens when you have improperly folded proteins
they become a clump of garbage and can be toxic. it causes many inherited human diseases. big protein-destroying strucutres, the proteasome, degrade the incorrect proteins
81
explain how proteasome and chaperones fight for protein
they both have a crush on fucked up proteins with exposed hydrophobic residues. they love it so much, and will compete for it. the longer it takes for a protein to fold, the more likely it is degraded
82
What is a proteasome, is it common. structure, and what does it act on
yeah its everyfuckingwhere, in the cytosol and nucleus. It's a hollow cylinder with caps on either end to prevent randomly eating shit, and a hollow cylinder in the middle. it eats proteins with ubiquitin chains. the longer the chain, the more likely it will be destroyed >:)
83
How is ubiquitin added onto proteins
Using ubiquitin conjugating system bade of E1, E2, E3.
84
What is E1
ATP dependent ubiquitin activating enzyme that yoinks a ubiquitin and puts it onto itself.
85
What is E2
Ubiquitin conjugating enzyme that yoinks uniquitin from E1 and works with E3
86
What is E3
binds to specific degradation sequences (degrons) in substrates. uniquitin is added onto a lysine residue on the target protein. process repeated to get polyubiquitination. ubiquitin does not touch E3.
87
What is a ubiquitin ligase? colloquially and officially
Sometimes E2-E3 are called ubiquitin ligase together, but E3 is the actual ubiquitin ligase.
88
How many of E1s, E2s and E3s do you have
a couple different E1s, about 30 different E2s and 100s of E3s.
89
What does monoubiquitination do
Monoubiquitination: histone regulation
90
What does multiubiquitination do, what is it
Multiubiquitination: endocytosis (like multiple on different spots)
91
What is polyubiquitination, what does it do
A chain of ubiquitin, linked through lysine 48 or 63 of the ubiquitin. for lysine 48, it causes proteosomal degradation. For lysine 63, it causes DNA repair.
92
How do ubiquitin ligases get activated
Phosphorylating E3 by protein kinase, allosteric transition caused by ligand binding on E3 or protein subunit addition
93
How do you activate a degradation signal which attracts ubiquitin ligases
Phosphorylation by protein kinase, unnmasking by protein dissociation, creation of destabilizing N-terminus
94
How can you post translationally regulate proteins
Make proteins on demand, ligand binding to activate, covalent modifications to activate, addition of subunit to activate, unmasking to activate, stimulation of nuclear entry, release from membrane
95
How does epinephrine activate pKA
epinephrine binds to GPCR, activating it, which goes through a g protein to make cAMP, which frees the two catalytic subunits from the regulatory subunits. The regulatory subunits bind to cAMP. the catalytic subunits promote breakdown of glucogen and inhibit glycogen synthesis. pKA promotes glucose release.
96
What does activated pKA do
it goes from the cytosol into the nucleus, which then activates genes with cAMP responsive elements (CRE), which are basically bits of DNA that CRE binding proteins (CREB) bind to. The CREB recruits CBP coactivator, which and the two bind to CRE, which activates the target gene. in the liver, this transcribes the gene glucose-6-phosphatase, which makes more glucose to be released into blood, and inhibits glycogen synthesis
97
How do proteins usually function
In large multi-protein complexes comprised of static and transient interactions.
98
What is an interactome map
complete collection of protein-protein interactions of an organism
99
What is a lysosome
an organelle where cell components no longer needed are degraded
100
How do post-translationally sorted proteins go to their destinations
Proteins going to mitochondria and plasmids go in unfolded, proteins going to nucleus and peroxisomes go in folded
101
How do proteins go into nucleus
Gates transport. The nuclear pore complexes (NPCs) are made of nucleoporins and let small things go freely but not DNA or proteins. The nuclear important receptor binds to nuclear localization signal on the protein, which is rich in lysine and argenine, which then binds to nucleoporins in the NPC.
102
How are nuclear export receptors related to nuclear import receptors
Structurally and evolutionally related.
103
Examples of exported cargo proteins
Proteins with nuclear export signals. newly assembled ribosomal subunits and RNA do too (because they all need to go to the cytoplasm!!!!!!!!!)
104
Explain the role of RAN-GTP in importing cargo proteins from the nucleus
Nuclear import receptor binds cargo in cytosol, then the two go to the nucleus. RAN-GTP binding causes cargo release in the nucleus. Then the empty import receptor and RAN-GTP go to the cytosol.
105
What does Ran-GAP do, where is it
GTP hydrolysis (changing GTP to GDP). Releases import receptor, export receptor, and relesae of cargo. It is in the cytosol
106
Explain the role of RAN-GTP in exporting cargo proteins from the nucleus
Nuclear export receptor binds Ran-GTP and cargo in the nucleus, then the three go to cytosol. Once GAP yeets cargo and export receptor off then the export receptor can go back to the nucleus
107
What does RAN-GEF do, where is it
It is a guanine nucleotide exchange factor in the nucleus, which promotes exchange of GDP for GTP by RAN (does the opposite of GAP)
108
How is NFAT an example of regulation of nuclear import and export
When there is a shit ton of calcium in the cell, protein phosphatase blocks nuclear export signal, which reveals the import one, and the protein goes in and activates gene transcription. This is because you want to get the calcium OUT. When theres little Ca2+ in the cell, ATP+ active protein kinase phosphorylate protein changes conformation to reveal the export signal and hasta la vista baby
109
How do proteins stay unfolded in the cytosol
By associating with Hsp70 chaperones
110
How do you get shit into the mitochondria
Nterminal amphipathic alpha helix guides proteins in (like the protein has an alpha helix). It binds to the import receptor, with a TOM on the outer membrane and a TIM on the inner membrane. The signal sequence in the membrane is then cleaved
111
How do you get shit into chloroplast
Also N terminal amphipathic alpha helix. Signals equence also cleaved. Outside membrane is TOC complex, inside is TIC. Sometimes, chopping off the signal sequence will reveal a hydrophobic thylakoid signal sequence
112
How do you get shit into peroxisome
3 amino acid at C terminal (SKL) that is not cleaved. The protein is folded (unlike for mitochondria and chloroplasts), and it is transported across by large translocator complex
113
How do proteins go to the ER
mRNA and ribosomes go to ER lumen. hydrophobic amino acids at N terminal is the ER signal sequence which first emerges from the ribosome. co-translational translocation occurs.
114
What does the ER do
synthesis and modification of proteins, synthesis of lipids
115
Where are some of the final destinations of proteins that go to the ER
soluble proteins, transmembrane proteins, proteins destined for the Golgi, secretion, lysosomes
116
What is a signal recognition particle (SRP)
a mix of protein and RNA that binds to the signal sequence with its hinge on the large ribosomal subunit. SRP receptor is on the membrane of the ER. both SRP and SRP receptors have GTP domains (that bind GTP). SRP and ribosome does not bind well to the receptor, you need an ER signal sequence as well
117
What cleaves the ER signal sequence
signal peptidases
118
How does the translocator prevent stuff from just leaving the ER
it is gated in a second direction
119
What are three ways singlepass proteins can be put in the membrane
The first two are basic, basically positive side faces the cytosol, can be either C or N terminal. The third one is that there is a signal sequence at the very tip of the N terminal which gets cleaved off, and the transmembrane domain is actually a stop transfer signal. COOH has to be in cytosol for this one
120
How are multipass proteins threaded into the membrane
The first TM domain is the internal start-transfer sequence, then the second TM domain is the stop-transfer sequence.
121
What are types of membrane proteins
Integral, lipid anchored, and peripheral
122
How are GPI-anchored proteins formed
has a c-terminal hydrophobic domain, which signals for GPI anchor while signal sequence is still in the membrane. The ER enzyme transfer protein to GPI anchor, and GPI anchored protein ends up on ER luminal side and can go to cell exterior
123
Where is glysolation common
In transmembrane and soluble proteins in the ER
124
What are the two types of glysosylation, which is more common
O-linked glycosylation, glucose on the oxygen of the side chain, which happens 10% of the time. N-linked glycosylation, on asparagine side chain's nitrogen, which happens 90% of the time.
125
How does N-linked glycosylation happen in the ER, what sequences can you put the oligosaccharide onto. Where does glycosylation happen
The N-linked oligosaccharide precursor is transferred by an oligosaccharyl transferase to an Asn on a protein being synthesized. This can be Asn-X-Ser or Asn-X-Thr. Proteins are only glysosylated on the ER lumen side.
126
What happens after you put the N-linked oligosaccharide onto a protein
3 glucoses removed, 1 mannose removed, then glycosylated protein is transported via vesicles to the golgi.
127
How is the golgi ordered, why is this important
From closest to furthest from ER: cis golgi network, cis cisterna, medial cisterna, trans cisterna, trans golgi network. Different places mean different modifications
128
Why do you need to glycosylate proteins
To mark where protein folds, to protect proteins on the cell surface from proteases. For cell adhesion, to allow formation of 3D structure.
129
What are some cargo proteins in vesicular transport
Transmembrane proteins, soluble proteins, some are bound by transmembrane cargo receptors
130
What do protein coats do for newly formed transport vesicles
Select cargo, give curvature to vesicle, promote vesicle budding
131
Where do COPI coated vesicles go
from Golgi to ER, between different Golgi cisternae.
132
Where do COPII coated vesicles go
from ER to Golgi (the normal direction)
133
Where do clathrin coated vesicles go
from Golgi and plasma membrane to endosome (sandwich)
134
What are the general steps in coat assembly and vesicle formation
GEF at the site of membrane yeets a gtp onto the Sar1 GTPase, then the Sar-1 GTP recruits coat proteins, which forms vesicle bud (like a little hill) and selects the cargo. The vesicle buds off, then it uncoats. This process for COPI and COPII proteins involve GAPs. Clathrin coated vesicles have a different mechanism. Now the vesicle can go to the target compartment
135
What GTPase do COPI and COPII vescicles use
COPI uses ARF. COPII uses Sar1
136
How do the GTPases in vescicle budding interact with the membrane
Once they get a GTP attached, they expose an amphipathic alpha helix which interacts with the membrane of the ER
137
What are the two layers on a vesicle
The inner layer binds to membrane and selects cargo. Outer layer associates with inner layer to promote formation of the coat. Coat proteins need to select cargo (transmembrane proteins), transmembrane cargo receptors (which bind soluble cargo proteins), and SNAREs.
138
How many subunits are in COPI coated vesicles, both inner and outer layers?
In the inner layer, there are 4 subunits, (β, γ, ζ, and delta)
139
How do uncoat a COPI coated vesicle specifically
y-COP binds to Arf GAP, which makes Arf-GTP go to Arf-GDP. Arf-GDP then detatches from the membrane
140
What are the subunits for the inner and outer layers of the COPII coated vesicles
Inner: 2 subunits (sec23/sec/24)
Outer: 2 subunits (sec13/sec31)
141
How to uncoat a COPII coated vesicle specifically
Sec23 has CAP activity, stimulated by sec 13 and sec31. Same GTP hydrolysis as in COPI, and Sar-GDP detaches from the membrane.
142
What are the subunits for clathrin coated vesicles
Inner: different adaptor proteins
Outer: clathrin (6 subunits)
143
How do you pinch off a clathrin coated vesicle
Dynamin, which has GTPas activity
144
What do you need to uncoat a clathrin coated vesicle
Hsp70 and auxillin
145
What is a triskelia
A fundamental unit in clathrin vesicles, which form a curved lattice that pinches off the membrane
146
Explain how docking/tethering proteins and fusion proteins fuse vesicles
GTP bound Rab-GTPase (on vesicle) binds Rab effector on target. Dock and tether, bringing vSNARE and tSNARE together. Helical domains coil, squishing out water. Cytosolic leaflets first fuse, leaving lumenal leaflets side by side
147
How do you dissociate SNARE complexes
NSF + adaptor proteins unwrap helical domains of SNAREs
148
What proteins have exit signals
Soluble proteins are bound by cargo receptors and transmembrane proteins are bound by COPII coat.
149
Do all cargo proteins in vesicles need exit signals
No, some are packaged because they are in high concentrations in the ER.
150
What moves vSNAREs and tSNAREs within vesicles so they can be used again
NSF
151
What happens when COPII coated vesicles fuse with each other
they form vesicular tubular clusters, which then go to the golgi
152
What brings escaped stuff back to the ER and how
COPI does, it brings back escaped ER resident proteins, proteins from ER involved in vesicle budding, these typically have ER retrieval signals. Soluble ER proteins ahve a KDEL signal and are bound by the KDEL receptor
153
What happens to the pH of the cell as you move towards the Golgi? how do KDEL receptors use this to bind and release
it gets lower and lower. At vesicular tubular clusters and golgi, the KDEL receptors will bind KDEL because acidic pH, and KDEL drops off at ER, which has a neutral pH
154
Why do some ER and golgi resident proteins not have retrieval signals
They have different transport rates from ER to golgi and vice versa. Proteins stuck in a certain compartment form big complexes and can't fit into vesicles.
155
How are transport vesicles kept close to the golgi cisternae
tethering proteins
156
What is the vesicular transport model
COPII coated vesicles move cargo from one golgi cisterna to the next. COPI return escaped resident ER proteins.
157
What is the cisternal maturation model
Cisternae all get promotions. vesicular tubular cluters fuse to become ER cis golgi network. Existing trans cisterna moves to trans golgi network. Retrograde transport by COPI vesicles
158
What is the TGN (trans golgi network)
a complex network of membranes and vesicles, a major branch point where proteins are sorted into different vesicles.
159
How are lysosomes formed
Vesicles with lysosomal hydrolases needed for lysosome function are transported to late endosome which mature into lysosomes.
160
How do acid hydrolases work, where are they made
Synthesized in the ER, processed in the Golgi. Degrade macromolecules, active at acidic pH. They starte off with n-linked oligosaccharide in the ER, phosphorylates mannose residue in cis-golgi network to form M6P., which receptors in the trans golgi network find, package and release into late endosome.
