Lecture 7 - Protein Sorting Ch. 15

Question 1

what does Cellular Compartmentalization allow for?

Answer 1

cellular specialization

Question 2

what dominates cellular mass?

Answer 2

Proteins, which almost all synthesis begins in the cytosol

Question 3

what is protein sorting?

Answer 3

The transfer of proteins from the cytosol to different organelles

Question 4

Three fundamentally different mechanisms by which
proteins move from one compartment to another?

Answer 4

1. Gated Transport
2. transmembrane transport using protein translocators
3. vesicular transport

Question 5

where are 5 areas proteins go to begin the “roadmap”

Answer 5

nucleus, mitochondria, endoplasmic reticulum, peroxisomes, plastids(chloroplast)

Question 6

where do all proteins eventually end up?

Answer 6

Cell exterior

Question 7

how do the cytosolic proteins “know”
where to go in the cell?

Answer 7

1. Proteins have specific amino acid sequences that direct them to their destinations.
2. Sorting signals are recognized by receptors that guide proteins to the right location.
3. Proteins without sorting signals stay in the cytosol.

Question 8

If a protein has a signal sequence on the N-terminus where will the protein go?

Answer 8

Import to the ER or the mitochondria

Question 9

What does recombinant DNA techniques do?

Answer 9

Between 2 proteins, it takes one of the signal sequence and it attaches it to the other protein. Both proteins will then go to their right location

Question 10

Signal sequences are both ____ and ____ to direct a protein to a specific destination

Answer 10

necessary and sufficient

Question 11

If a protein has a signal sequence on the C-terminus where will the protein go?

Answer 11

import into peroxisome or retention in lumen of ER

Question 12

If a protein doesn’t have a signal sequence where will the protein go?

Answer 12

It will stay in the cytosol

Question 13

Why do eukaryotic cells require a separate compartments?

Answer 13

a strategy to confine different metabolic processes and the proteins which are required for them for cellular specialization

Question 14

Where can sorting signals be found in the amino acid sequence?

Answer 14

N-term or C-term

Question 15

what usually happens to finished proteins?

Answer 15

The N-term is usually removed; other signal sequences become a part of the mature protein

Question 16

Why must a sorting signal be distinct?

Answer 16

To allow selective recognition by the appropriate sorting receptor.

Question 17

what is the first to be synthesized on polypeptide chain?

Answer 17

The N-term where synthesizes goes N -> C direction

Question 18

where in the amino acid sequence of a protein is the signal sequence for import/export of the nucleus located?

Answer 18

internally within the amino acid (no N or C term)

Question 19

What is KDEL, and what does it do?

Answer 19

KDEL is an ER retention signal, keeping proteins in the endoplasmic reticulum. It is named after its single-letter amino acid sequence: Lys-Asp-Glu-Leu

Question 20

What are the internal signal sequences?

Answer 20

signal sequence is used to start the protein transfer, called a start-transfer sequence, is never removed from the polypeptide as an anchor

Question 21

What type of sorting signals direct protein translocation into organelles?

Answer 21

Linear signal sequences guide proteins into organelles.

Question 22

What is gated transport, and where does it occur?

Answer 22

A transport mechanism where molecules move between the nucleus and cytosol through nuclear pores.

Question 23

What is the function of the lamin meshwork?

Answer 23

It provides structural support and serves as an anchoring site for nuclear pore complexes and chromosomes.

Question 24

What is the composition of the nuclear pore complex (NPC) in animal cells?

Answer 24

molecular mass of 125 million Da, consists of 30 different nucleoporins in multiple copies, and contains a water-filled channel

Question 25

How many NPCs does a typical mammalian cell have, and how do they function?

Answer 25

The nuclear envelope has 3000-4000 NPCs, transporting 1000 macromolecules per second in both directions simultaneously. NPCs are constitutively open.

Question 26

Is nuclear import/export selective?

Answer 26

Yes, Import: Histones, DNA polymerases, transcriptional regulators. Export: mRNA, rRNA, tRNA, and other RNA molecules.

Question 27

What is the size limit for passive diffusion through the NPC?

Answer 27

Diameter is ~40nm, allows small molecules (<5,000 Da) to diffuse freely. Molecules >40,000 Da require sorting signals for active transport. Larger molecules, like ribosomal subunits and some viruses, can pass through with assistance.

Question 28

How do nuclear import receptors function?

Answer 28

Soluble cytosolic proteins that bind both the NLS on cargo proteins and NPC fibrils to mediate transport. Different receptors recognize different NLS signals. Nuclear proteins are transported fully folded, unlike ER/mitochondrial transport. Similar export receptors handle mRNA transport.

Question 29

what protein triggers GTP hydrolysis located only in the cytosol?

Answer 29

Ran-GAP (GTPase-activating protein), converts Ran-GTP to Ran-GDP

Question 30

What is Ran-GEF, and where is it located?

Answer 30

accessory protein Ran-GEF (guanine nucleotide exchange factor) is found only in nucleus & facilitates exchange of GDP for GTP on Ran (not substrate-level phosphorylation). This creates a higher concentration of Ran-GTP in the nucleus.

Question 31

what is NLS?

Answer 31

nuclear localization signal

Question 32

what are the 2 conformations of GTPase Ran?

Answer 32

One carrying GTP and the other carrying GDP

Question 33

What is the kinase that phosphorylates Ran-GDP to Ran-GTP?

Answer 33

Ran does not undergo phosphorylation, conversion is done by GEF which exchanges GDP for GTP, which replaces the entire nucleoside

Question 34

how do proteins transport to mitochondria and ER?

Answer 34

from cytosol through protein translocators (located in membrane) that must be unfolded

Question 35

the matrix of the mitochondria?

Answer 35

high concentration of enzymes, fatty acid oxidation and citric acid cycle

Question 36

Inner membrane of mitochondria?

Answer 36

folded cristae carries out oxidative phosphorylation. non permeable, requires transport across membrane

Question 37

outer membrane of mitochondria?

Answer 37

contains porins (large channel-forming) render mitos permeable to molecules

Question 38

intermembrane space of mitochondria?

Answer 38

site of proton buildup that drives ATP synthesis

Question 39

how is mitochondria DNA synthesized?

Answer 39

encoded in the cell nucleus and imported from the cytosol

Question 40

How do new mitochondria form and how are mitochondrial precursor proteins kept unfolded in the cytosol?

Answer 40

produced by the growth and fission of preexisting mitochondria, relying on protein import for expansion. Mitochondrial precursor proteins remain unfolded in the cytosol w/ the help of chaperone proteins. These include HSP70 family chaperones and proteins that bind directly to mitochondrial signal sequences to prevent premature folding.

Question 41

chaperones aid in what?

Answer 41

helps fold proteins that are misfolded or newly synthesized.

Question 42

mitochondrial proteins are first fully synthesized as....

Answer 42

precursor proteins in the cytosol

Question 43

signal sequences that direct proteins into mitochondria has potential to form....

Answer 43

amphiphilic alpha helix

Question 44

mitochondrial precursor proteins are imported as.....

Answer 44

unfolded polypeptide chains

Question 45

how does the translocation into mitochondria work?

Answer 45

n-terminal sequence is recognized by transmembrane receptors, which is translocated through translocator through both membranes, signal sequence is cleaved and degrades in the matrix space

Question 46

what occupies 10% of the total cell volume?

Answer 46

ER lumen and continuous sheet to the nuclear membrane

Question 47

ER serves as....

Answer 47

entry point for proteins destined for other organelles

Question 48

The ER membrane is the production site of all transmembrane proteins and lipids that include?

Answer 48

ER itself, Golgi, lysosomes, endosomes, secretary vesicles, and plasma membrane

Question 49

majority of proteins that will be secreted to cell exterior...

Answer 49

are initially delivered to the ER lumen

Question 50

what are the top 4 things that make up cell volume?

Answer 50

cytosol (54%), mitochondria (22%), ER (12%), Nucleus (6%)

Question 51

what is the co-translational translocation process?

Answer 51

ribosome that is directly attached to ER membrane which enables one end of the protein to be translocated into ER while protein is still being assembled

Question 52

what is post-translational translocation?

Answer 52

import of proteins into mitochondria and peroxisomes after synthesis of protein after translocation

Question 53

what do free ribosomes in cytosol do?

Answer 53

allows for the mRNA encoding with no ER signal sequence, which remains in the cytosol

Question 54

what do membrane-bound ribosomes attached to the ER do?

Answer 54

Has a ER signal sequence which directs the ribosome to the ER membrane where the protein is fed into the mitochondria

Question 55

What 2 components is needed to direct ribosome to the ER membrane?

Answer 55

Signal recognition particle (SRP) in the cytosol and SRP receptor embedded in the ER membrane

Question 56

what two proteins are synthesized by membrane-bound ribosomes?

Answer 56

water-soluble proteins are completely translocated across the ER membrane and prospective transmembrane proteins which are partly translocated across the membrane and become embedded into it

Question 57

how does the water soluble protein translocate across the ER membrane

Answer 57

translocator binds to the signal sequence, where is cleaved by signal peptidase. This is ejected into bilayer where it degrades. The protein translocator closes once the polypeptide is release in the ER lumen.

Question 58

proteins that remain in the ER have a what?

Answer 58

C-terminus KDEL signal sequence

Question 59

how does prospective transmembrane proteins translocate across the ER membrane?