Chapter 13 - Intracellular Compartments and Protein Transport

What is Biomolecular Condensate

What is Biomolecular Condensate

membraneless organelles/sub compartments

Biomolecular condensates often contain

• RNA in addition to protein

• perform a particular function

What is the largest biomolecular condensate

The nucleolus

An animal cell contains a basic set of__________

membrane enclosed organelles

What does each membrane compartment contain

a unique set of molecules that carry out a specialized function

Contains many metabolic pathways, protein synthesis, the cytoskeleton

Cytosol

Contains main genome, DNA and RNA synthesis

Nucleus

Synthesis of most lipids, synthesis of proteins for distribution to many organelles and to the plasma membrane

Endoplasmic Reticulum

Modification, sorting, and packaging of proteins and lipids for either secretion or delivery to another organelle

Golgi apparatus

ATP sythesis by oxidative phosphorylation

Mitochondria

Sorting of endocytosed material

Endosomes

Intracellular degradation

Lysosomes

Oxidative breakdown of toxic molecules

Peroxisomes

Surrounded by double membrane which consists of a nuclear envelope and nuclear pores

Nucleus

True or false, the outer nuclear membrane is continuous with the membrane of the ER

True

Types of ER

• Smooth

• Rough

ER interior is also called

ER lumen

Receives lipids and proteins from ER, modifies them and dispatched to other destinations. Also the production of vesicles

Golgi Apparatus

This compartment is surrounded by a db membrane, with is also a site for oxidative phosphorylation

Mitochondria

Digestive enzymes that degrade worn out organelles, macromolecules and particles taken into cell by endocytosis

Lysosomes

Series of compartments involved in sorting of ingested molecules. Some recycle back to PM, others deliver to lysosomes

endosomes

Enzymes that brwak down lipids and destroy toxic molecules

peroxisomes

Membrane enclosed organelles on average occupy nearly ______ of the volume of a cell

1/2

The area of the ER membrane is _________ than the ______ in a typical mammalian cell

• 20-30x greater

• PM

True or False: In terms of area and mass, the PM is only a major membrane in most eukaryotic cells

False

To break open cells and tissues you will need

• Homogenate

• Extract

• Lysate

How do we isolate cellular organelles when breaking open cells and tissues

differential centrifugation

differential centrifugation separates components based on

size and density

During the evolution of membrane enclosed organelles, what compartment remained isolated

Mito genome

Why is continuous protein synthesis and delivery necessary

• Proper functioning of cells

• For any cell to grow and divide

Newly synthesized proteins must be accurately delivered to their ____________

appropriate organelle

Direct delivery of proteins from cytosol lead into

• Interior of the nucleus

• mitochondria

• the chloroplast

Indirect delivery of proteins via the ER leads to

• Inner nuclear membrane

• Golgi

• lysosome

• endosome

Where does protein synthesis begin

On ribosomes in the cytosol

Mitochondrial and Chloroplast protein facts

• few mitochondrial and chloroplast proteins that are synthesized on ribosomes inside these organelles

• most mitochondrial and chloroplast proteins are made in the cytosol and subsequently imported

protein destination after its synthesis in the cytosol is dictated by

its amino acid sequence

directs the protein to the organelle in which it is required

a sorting signal

What happens to proteins that lack a sorting signal

proteins remain permanent residents of the cytosol

Can a membrane-enclosed organelle import a water-soluble protein to its interior?

it really depends on the organelle

Ways to transport a water-soluble protein in a membrane-enclosed organelle

• Nuclear pore

• Protein translocators

• Transport vesicles

what is a signal sequence

a sequence that directs proteins to the correct destination

In specifying the same destination, properties such as hydrophobicity and placement of charged aa appear to be more import than the aa sequence itself, True or False?

True

True or False: Properties are often not removed from the finished protein

False

____________ encloses the nuclear DNA and defines the nuclear compartment; is formed from two______________

• Nuclear envelope

• Concentric membranes

Inner nuclear membrane contains proteins that act as a binding site for

chromosomes

Inner nuclear membrane contains proteins that act provide anchorage for

The nuclear lamina

Finely woven meshwork of protein filaments that lines the inner face of this membrane and provides structural support to the nuclear envelope

Nuclear lamina

what is that nuclear lamina apart of

Inner nuclear membrane

closely resembles the membrane of the ER, with which it is continuous

Outer nuclear membrane

Outer nuclear membrane is apart of

the nuclear envelope

The nuclear envelope is perforated by

Nuclear Pores

large, elaborate structure composed of a complex ~30 different proteins, each present in multiple copies

Nuclear pores

Nuclear pores are lined with

Extensive unstructured disordered regions of polypeptide chains

What do proteins to be delivered inside the nucleus require

Nuclear localization signals

What do Nuclear localization sites consist of?

one or two short sequences containing several positively charged Lys or Arg

What are nuclear localization sites usually recognized by?

cytosolic proteins called nuclear import receptors

Directionality of nuclear import receptors is guided by what?

• the hydrolysis of nucleoside triphosphate,

• GTP, by monomeric GTPase named Ran

The differently localized accessory proteins that Ran converts from one form to another are

• Ran - GDP - mainly in cytosol

• Ran - GTP - high concentration in the nucleus

• Ran - GAP - GTPase-activating protein

• Ran-GEF - Guanine nucleotide exchange factor

What drives protein and RNA traffic from the nucleus to the cytosol

Nuclear export receptors

What recognizes specific nuclear export signals

Nuclear export receptors

What also uses Ran to couple the transport to an energy source

Nuclear export receptors

How are proteins transported into nucleus

in their fully formed conformation

For delivery into the nucleus, nuclear pore proteins rapidly usher macromolecules in both directions through each pore

What usually has a signal sequence at their N-terminus and must cross both, the inner and the outer membranes that surround each of these organelles

Proteins to be delivered into the mitochondria

Where does translocation take place

At specialized sites where the two membranes are held closely together

Proteins being delivered to the mitochondria are _________ as it is transported

Unfolded

After proteins are delivered to the mitochondria, _______________ is removed after translocation is complete

Signal sequencing

proteins inside mito help to pull the protein across the two membranes and refold it once it is inside

Chaperone proteins

How do proteins enter peroxisomes

both the cytosol and ER

a short sequence of only 3 aa serves as an import signal peroxisomes acquire the bulk of their proteins via selective transport from

the cytosol

few proteins arrive via vesicles that bud from the ER true or false

True

Entry point for proteins designed for ER and other organelles

• Golgi

• Endosomes

• Lysosomes

• Cell surface

One proteins enter the ER, they will not re-enter what?

The cytosol

How do proteins that enter the ER while being synthesized reach their destination

by transport vesicles

Proteins translocated into ER

• Water-soluble proteins

• TM proteins

completely translocated across the ER membrane and release into ER lumen

Water-soluble proteins

only partially translocated across the ER membrane and become embedded in it

TM proteins

ER signal sequence consists of

• a segment of 8 or more hydrophobic aa

• also involved in the process of translocation across the membrane

True or False: proteins are threaded across the ER membrane before the polypeptide chain has been completely synthesize

True

True or False: ribosome synthesizing the protein does not have to be attached to the ER membrane

False

How do free and bound ribosomes differ

The proteins they make

What happens when a ribosome is making a protein with an ER signal sequence

The signal sequence directs the ribosome to the ER membrane

proteins with an ER signal are translocated as they are being made, additional energy is required for their import: True or False

False

proteins with an ER signal are translocated as they are being made, no additional energy is required for their import, this is because

the elongation of each polypeptide provides the thrust needed to push the growing chain through the ER membrane

Signal-recognition particles bind to both?

The exposed ER signal sequence and the ribosome

What does the SRP receptor do?

• Recognizes the SRP embedded in the ER membrane

One SRP binds to SRP receptors what happens?

• SRP is released

• receptor passes on the ribosome to a protein translocator in the ER membrane

When an ER signal and an SRP direct a ribosome to the ER membrane, what happens to the growing polypeptide

it is threaded across the ER membrane through a channel in the translocator

An ER signal sequence for soluble proteins

• is almost always at N-terminus

• functions to open the translocator

  • remains bound to the translocator

  • removed by TM signal peptidase

What is a single pass transmembrane protein

TM protein with a single alpha-helical membrane- spanning segment

Describe the single pass transmembrane protein process

• N-term signal sequence initiates translocation

• process is halted by additional sequence of hydrophobic aa, stop-transfer sequence

What do db pass membrane proteins do

Start-transfer sequence

What are multi-pass transmembrane proteins

when many hydrophobic alpha helices span the bilayer

Name the properties of multipass TM proteins

• additional pairs of start- and stop-sequences

• even # of transmembrane alpha helices

• both N- and C-terminus oriented toward the same side of the membrane

• odd # of transmembrane alpha helices

• opposite orientation of N- and C-terminus

________________ direct insertion and orientation of various classes of integral proteins into ER membranes

Topogenic sequences

_________________ requires not only import of new proteins, but also incorporation of new lipids into the organelle membranes

Growth and maintenance of organelles

phospholipids are manufactured

by enzymes bound to the cytosolic surface of ER

True or False: organelles that are part of endomembrane system can receive lipids via transport vesicles

True