Sorting Flashcards
1
Q
plasma membrane
A
outer boundary of cells
phospholipid bilayer
protection, transporters, cell signaling
2
Q
nucleus
A
houses genome
RNA and DNA synthesis
3
Q
cytoplasm
A
cytosol + cytoplasmic organelles
intermediary metabolism
4
Q
endoplasmic reticulum
A
with ribosomes = rough
w/o = smooth
protein/lipid synthesis, protein folding, quality control, Ca storage, signaling
5
Q
golgi apparatus
A
stacks of disc-like compartments
post-translational changes to proteins/lipids
trafficking
6
Q
mitochondria
A
outer and inner membrane matrix
powerhouse of cell
signaling
cell differentiation and death
7
Q
what happens when a mitochondria is leaky?
A
apoptosis
8
Q
lysosomes
A
contain digestive enzymes to degrade organelles and biomolecules
9
Q
peroxisomes
A
small vesicular compartments that contain enzymes used in oxidative rxns
10
Q
topological compartments
A
- nucleus and cytosol
- secretory and endocytic organelles
- mitochondria
11
Q
3 types of transport for trafficking
A
- gated transportation
- TM transportation
- vesicular trafficking
12
Q
gated transportation
A
between nucleus and cytosol
thru nuclear pores
bidirectional
13
Q
TM transportation
A
cytosol to peroxisomes, plastids, mito, ER
monodirectional
membrane transporters directly transport proteins from cytosol to target
14
Q
vesicular trafficking
A
ER —-> elsewhere
use of membrane bound vesicles to transport molecules
15
Q
what guides protein sorting?
A
protein sorting signals
16
Q
protein sorting signals
A
sequence of AAs on protein
can be anywhere or multiple places in protein
but when folded they come together to form a signal patch
necessary and sufficient
17
Q
signal peptidase
A
after a protein has reached it’s final destination
peptidase can cleave the signal sequence off because it is no longer needed
18
Q
what is more important in a signal sequence?
A
physical properties are more important than the actual sequence
19
Q
_____ receptors recognize and read signal sequences.
A
complimentary receptors
20
Q
import into nucleus
A
lys and Arg rich
21
Q
import into mitochondria
A
combination of + charged and hydrophobic AAs
22
Q
import into ER
A
bunch of hydrophobic AAs
23
Q
export from ER
A
KDEL
Lys-Asp-Glu-Leu-COO-
24
Q
what molecules are exported from the nucleus?
A
mRNA and tRNA
25
NPC
nuclear pore complexes
26
NPC characteristics
composed of nucleoporins
octagonal
extending fibrils facilitate mvt
3000-4000 NPC per nucleus
27
NPC components
```
cytosolic fibers
scaffold nucleoporins
membrane ring proteins
channel nucleoporins
disordered region of C nucleoporins
nuclear basket
```
28
NLS
nuclear localization signals
direct mvt of protein into nucleus
rich in + AAs (lys Arg)
located in loops or patches on surface of cargo
29
NIR
nuclear import receptors
cytosolic proteins that:
recognize NLS and bind to it and NPC proteins
30
types of NIR binding
direct binding
indirect binding
via adaptor protein
31
NPC binding sites for NIR
FG repeats
| phenylalanine glycine repeats
32
after NIR delivers protein to destination where does it go?
delivers protein to nucleus and returns to cytosol
33
NES
nuclear export signals
same as import just signal opposite direction
34
NER
nuclear export receptors
complimentary to NES, bind to it and NPC proteins to move out of the nucleus
35
monomeric G protein
Ran
36
cytosolic Ran vs. nuclear Ran
GDP cytosol
GTP nucleus
37
Ran cytosol
GAP
GAPase activating protein
cleaves phosphate bond to keep Ran as GDP
38
Ran nucleus
GEF
guanine exchange factor
exchanges guanine with a GTP guanine
does not add Pi group
39
driving factor for gated transportation
Ran-location type gradient
40
Ran-GTP binding......
```
binds to (NIR + cargo)
cargo is released
NIR--Ran exit nucleus
GAP cleaves
Ran-GDP now ready for another cycle
```
41
proteins that contain both NLS and NES sequences
shuttling proteins
42
relative rate of gated transportation controls....
homeostasis = importing = exporting
import greater = nuclear
export greater = cytosolic
43
what controls transportation?
genes
keep proteins out of nucleus until they are needed
44
how is transportation controlled?
by turning NLS/NES on or off
45
mechanisms for transportation control?
phosphorylation
proteolysis
binding to inhibitory proteins
46
two examples given in this lecture: gated transportation
T-Cell activation
| low cholesterol
47
mitochondria structures
outer memb.
intermemb. space
inner memb.
inner matrix
cristae of inner memb.
48
significance of cristae
```
folds in inner membrane
function to increase surface area
```
49
source of mitochondria protein
most are encoded by nuclear DNA
| but some are made in the mito itself, has it's own translation machinery
50
translocation
mvt of proteins of membrane
51
what directs proteins to a specific organelle or compartment?
signal sequences
52
mitochondria signal sequences
located at N terminus or in middle
| amphiphilic alpha helix shape
53
describe amphiphilic alpha helix
shape of proteins destined for mitochondria
created by positive residues on one end and hydrophobic ones on other end
54
nonpolar residues are ?
hydrophobic
55
how do receptor proteins recognize precursor proteins bound for mitochondria?
by the alpha helix configuration not the signal sequence itself
56
where are protein translocators located?
on the membrane of organelle protein is imported to
57
protein translocators of mitochondria
multi-subunit protein complexes that mediate translocation
58
list the mitochondrial translocators
cytosol to inter space
TOM
SAM
inter space to matrix
TIM 22
TIM 23
OXA
59
translocator that transfers or inserts all proteins from cytosol to outer membrane
TOM
60
sorting and assembly machinery
SAM
| Translocates and inserts or folds beta barrel proteins
61
mediates insertion of specific subclass proteins
TIM22
| ATP, ADP, Pi transporter
62
transport of soluble proteins and insertion of proteins into inner membrane
TIM23
63
insertion of proteins synthesized in the mitochondria
OXA
a few exceptions recently found
64
TOM and TIM have 2 components
receptors for precursor protein
translocation channel
65
describe a precursor protein
unfolded proteins in cytosol
| maintained by chaperone hsp70
66
describe protein import through TOM
TOM binds to signal seq.
chaperones are stripped off --ATP
protein fed thru channel into space
peptidase cleaves signal
67
when is energy required in TM transport?
to remove hsp70
to remove mito hsp70 in matrix
for hsp60 to fold/refold protein in matrix
68
what drives protein mvt through TOM?
ATP hydrolysis drives removal of hsp70
| free unfolded peptide is then pulled thru TOM
69
what drives protein mvt through TIM?
the electrochemical membrane potential gradient, drives the positive protein by electrophoresis
protein wants to get to the negatively charged matrix
70
mitochondrial hsp70
binds to protein in matrix and helps pull it thru TIM23
ATP is required for hsp70 to release
71
hsp60
binds to protein in mito matrix and helps fold the imported protein
requires ATP to do so
72
integration within the outer mitochondrial membrane
pass thru TOM
chaperones bind
protein binds to SAM
SAM - inserts and folds protein into memb.
73
example of TM proteins on outer mitochondrial membrane
porins
74
ER structure
network of branching tubules and sacs
membrane is continuous with nuclear memb.
internal space = ER lumen
75
types of ER translocation
co-translational
post-translational
76
co-translational translocation
mvt into ER
ribosome still attached
translation still in process
77
post-translational translocation
mvt into ER
| translation finished
78
ER signal sequence
AA specific order varies
8 or more nonpolar/hydrophobic AA's in center or protein
79
signal sequence guidance to ER---2 factors
SRP
| SRP receptor
80
what does SRP stand for?
signal receptor particle
81
SRP structure
6 proteins bound to a small RNA backbone
rod shaped
large hydrophobic pocket
82
describe the hydrophobic pocket of SRPs
lined by methionines
| accommodates hydrophobic signal seq. of varying size, shape, sequence
83
SRPs cycle back and forth between ?
cytosol and surface of ER membrane
84
what do SRPs bind to?
ER signal sequence on protein
SRP receptor of ER membrane
in co-translational: to large unit of ribosome
85
where does a SRP bind to a ribosome?
to the large unit of ribosome at the elongation factor
and binds to the ER signal sequence of the protein being translated
86
describe steps of co-translational translocation
```
SRP binds ribosome
SRP binds ER signal seq.
translation paused
travel to ER
bind to SRP receptor
translation restarts
protein fed thru translocator
SRP recycled
```
87
where are the SRP receptors located?
next to translocators on ER membrane
88
describe ER translocators
circular shape
3 subunits
largest surrounds pore
central pore
89
describe ER translocator pore
water filled
core = Sec61 complex
gated by short helix
opens and closes as needed
90
what are the 2 states of the ER translocator?
open --- full circle, pore plugged
closed --- 3/4 circle shape, pore plug displaced
signal seq. bound in open 1/4
91
signal sequence that interacts with a specific site within the pore
start-transfer signal
also interacts with lipid components of ER membrane. acts as dual recognition to ensure specificity
92
what does the start-transfer signal do?
activates the pore opening
allowing entry to lumen
peptidase cleaves it off
93
integration of TM proteins to ER - requirements
some portion of the protein must pass thru the translocator before a stop-transfer signal is reached
94
what initiates integration of TM proteins in the ER?
N terminus
95
describe a stop transfer signal
a hydrophobic region in the polypeptide that stops translocation
peptidase cannot cleave because it becomes integrated into bilayer via the lateral gate of the translocator
96
can a TM protein be multi-integrated?
yes, single or multiple
depends upon the combination of start and stop transfer signals
97
in an ER TM protein which side of the membrane does each terminus of the protein end up on?
either side
they both can exist on the same side too
98
ER integration predictions
we can utilize software to predict the amount of integration of a protein
based upon the physical properties of the sequence itself
