Lecture 16 - Protein sorting at the cytoplasm

Question 1

How does the cell know where a protein is supposed to go?

Answer 1

linear signal sequences are the sorting signals for protein translocation into organelles

Question 2

Nuclear envelope

Answer 2

consists of 2 concentric membranes that are perforated by nuclear pore complexes

Question 3

Nuclear pore complexes

Answer 3

perforate the nuclear envelope in all eukaryotes
- composed of roughly 30 different proteins, or nucleoporins (mediate the passage of molecules)

Question 4

Mesh of the nuclear pore complex

Answer 4

formed by unstructured proteins; acts as a sieve that restricts the diffusion of large macromolecules while allowing smaller molecules to pass

Question 5

Nuclear localization sequence (NLS)

Answer 5

proteins with this sequence are directed into the nucleus
- nuclear localization signals have flanking basic clusters

Question 6

What sequence do proteins need to leave the nucleus?

Answer 6

a nuclear export sequence (NES)

Question 7

Import receptors (importins)

Answer 7

soluble cytosolic proteins that contain multiple low affinity binding sites for the FG repeats found in the unstructured domains of several nucleoporins

• they bind to proteins with NLS and mediate passage into nucleus

Question 8

What does the importin-cargo complex do?

Answer 8

it locally dissolves the gel-like mesh and can diffuse into and within the NPC pore

Question 9

What is the role of Ran?

Answer 9

the concentration of Ran bound to GTP provides energy and directionality
- for importins, Ran-GTP in the nucelus promotes cargo dissociation

Question 10

Exportins

Answer 10

NES receptors
- Ran-GTP promotes cargo BINDING

Question 11

GAP and GEF

Answer 11

GAP = GTPase-activating protein
- triggers GTP hydrolysis (GTP => GDP)
- cytosolic

GEF = guanine nucleotide exchange factor
- promotes the exchange of GDP for GTP
- nuclear

Question 12

Alpha and beta subunits of importins

Answer 12

alpha: binds the nuclear localization signal (NLS)
- no NLS => alpha subunit cannot bind => protein stays in cytoplasm

beta: binds the unstructured chains

Question 13

How does the importin-beta carry cargo inside?

Answer 13

in the cytoplasm a GTP molecule is hydrolyzed and Ran dissociates (releasing cargo molecule)

Question 14

What do exportins bind to?

Answer 14

1. the export signal (on cargo) either directly or via an adaptor
2. NPC proteins to guide their cargo into the cytosol

Question 15

Can proteins contain both NESs and NLSs?

Answer 15

yes!

proteins can constantly be shuttled between the nucleus and the cytosol

Question 16

Where are most mitochondrial proteins synthesized?

Answer 16

in the cytoplasm and then translocated into the mitochondria

Question 17

Do mitochondrial and plastid proteins have signal sequences?

Answer 17

yes; they must be located at the N-terminus part b/c the pores of mitochondria and plastids are small so only unfolded proteins can fit through

Question 18

TIM & TOM systems

Answer 18

move polypeptide chains through the 2 membranes of the mitochondria

TOM = translocase of the outer membrane
TIM = translocase of the inner membrane

TIC and TOC for chloroplasts

Question 19

Hsp-70 chaperones

Answer 19

interact with mitochondrial proteins at the cytoplasm and prevent them from folding and aggregating