Chapter 11 - Protein Sorting

network of membrane-enclosed tubules and sacs that extends from the nuclear membrane

endoplasmic reticulum

organelles involved in sorting and targeting of proteins

ER, Golgi apparatus, endosomes, lysosomes

secretory pathway for proteins

rough ER → Golgi → secretory vesicles → outside of the cell

destination for proteins synthesized on free ribosomes

cytosol, nucleus, or other organelles

destination for proteins synthesized on membrane-bound ribosomes

directly into the ER through translocon

proteins move into the ER during their synthesis on membrane-bound ribosomes

cotranslational translocation

proteins move into the ER after translation has been completed on free ribosomes

posttranslational translocation

ribosomes are targeted to the ER by a signal sequence

cotranslational pathway

method of removing signal sequences from secretory proteins

proteolytic cleavage

length of signal sequences

~20 amino acids

stretch of hydrophobic residues located at the amino terminus of the polypeptide chain

signal sequence

recognizes the signal sequence and escorts the complex to the ER membrane

signal recognition particle (SRP)

enzyme that cleaves the signal sequence

signal peptidase

location of signal peptidase

inside the ER lumen

proteins destined for incorporation into membranes are:

initially inserted into the ER membrane

structure of the membrane-spanning region of membrane proteins

α-helical with hydrophobic amino acids

number of times the α-helical region can cross the membrane

once

when inserted, membrane proteins have a smaller domain in this location

cytoplasm

sequence in the middle of the protein that halts translocation and inserts the protein into the ER membrane

transmembrane sequence (α-helical)

sequence that is recognized by the signal processing particle (SRP) but is not cleaved by signal peptidase

transmembrane sequence (α-helical)

allows a protein to span the ER membrane multiple times

series of transmembrane sequences with alternating orientations

assist in folding and assembly of newly translocated polypeptides

role of lumenal ER proteins

binds unfolded polypeptide chains as they cross the membrane, then mediates folding and assembly of multi-subunit proteins

Hsp70 chaperone BiP

locations where protein folding and processing occurs

within the ER lumen or across the ER membrane (during translocation)

reducing environment

cytosol

oxidizing environment

ER interior

protein sequence where N-linked glycosylation occurs

asparagine residues

function of glycosylation

prevents protein aggregation in the ER, provides signals for sorting

function of disulfide bond formation

helps form the 3D secondary structure in protein folding

glycolipids that allow the attachment of a new peripheral protein to the membrane

GPI anchor

GPI anchors are added to this section of a polypeptide

the carboxy terminus

screens new proteins and removes them from the ER if they are misfolded or incorrect

ER-associated degradation (ERAD)

ERAD process

misfolded proteins are identified by calreticulin, targeted back to the cytosol through ubiquitin ligase complex, and degraded by the proteasome (or, protein is modified and re-tested)

protein folding sensor

calreticulin

location where the glycoprotein is ubiquitylated

cytosol

activated when an excess of unfolded proteins accumulates

unfolded protein response (UPR)

unfolded protein response (UPR) effect on chaperone production

increased production

unfolded protein response effect on protein production

decreased production

UPR process

accumulation of unfolded proteins leads to ER expansion and increased chaperone production. If protein folding isn’t adjusted to a normal level, the cell undergoes programmed cell death

IRE1 cleaves pre-mRNA of XBP1, activating it to stimulate transcription of UPR genes in the nucleus

first pathway to promote chaperone synthesis

ATF6 is cleaved to release the active ATF6 transcription factor

second pathway to promote chaperone synthesis

PERK inactivates eIF2 through phosphorylation to inhibit translation and reduce amount of protein entering the ER

pathway to decrease protein synthesis

lipid production

main function of the smooth ER

phospholipids, glycolipids, cholesterol

lipid types that make up the eukaryotic membrane

location where phospholipids are synthesized and added

cytosol side of the ER membrane

difference between sphingomyelin and other phospholipids

uses serine instead of glycerol

enzyme that transfers phospholipids through the ER membrane to the inner layer

flippases

site of ceramide synthesis

endoplasmic reticulum

site where ceramide is converted to glycolipids or sphingomyelin

Golgi apparatus

a lipid that is a source for two other lipids

ceramide

type of ER abundant in cells with active lipid metabolism

smooth ER

organ with a lot of smooth ER

liver

exported from the ER in vesicles that fuse to form the ERGIC, and then are released into the Golgi

method of transporting proteins and phospholipids

formed by vesicles from the ER fusing together

ER-Golgi intermediate compartment (ERGIC)

direction in which proteins and lipids are transported

towards the Golgi and occasionally in reverse back to the ER

targeting sequence at the carboxy terminus that directs proteins back to the ER

KDEL and KKXX

cisternae (flattened membrane-enclosed sacs) and associated vesicles

structures of the Golgi apparatus

location where proteins enter the Golgi

convex cis face

location where proteins are transported through and leave the Golgi

concave trans face

locations where the Golgi transports proteins to

endosomes, lysosomes, plasma membrane, secretion

compartments of the Golgi apparatus

cis, medial, trans compartments, and trans-Golgi network

receives molecules from the ERGIC

cis compartment

location in the Golgi where most modifications are done

medial and trans compartments

sorting and distribution center

trans-Golgi network

proteins are carried in the cis to trans direction in this Golgi structure

cisternae

added in the ER, modified in the Golgi by a sequence of reactions in different compartments

N-linked oligosaccharides

carbohydrates are added to side chains of serine and threonine (sugars are added to a core protein)

O-linked glycosylation

structure formed by sugar molecules added to a core protein

dimer

routes for molecule transport from the Golgi to the cell surface

direct transport, recycling endosomes, regulated secretory pathways

protein with three arms that form interlaced complexes

clathrin

families of vesicle coat proteins

COPII-coated, COPI-coated, clathrin-coated

carry proteins forward from the ER to the Golgi

function of COPII-coated vesicles

carry proteins back to the ER from the Golgi

function of COPI-coated vesicles

carry proteins back and forwards between the Golgi, endosomes, lysosomes, and plasma membrane

function of clathrin-coated vesicles

GTP-binding proteins that regulate coated vesicle formation

ARF1 and Sar1

critical for fusion of vesicle and target membrane

tethering factors and RAB

transmembrane proteins that facilitate vesicle and membrane pairing

SNAREs (vesicle SNARE and target SNARE)