S16-S17

S16: Protein modifications; S17: protein trafficking

Phosphorylation of proteins occurs at which 3 AAs?

Ser, Thr, Tyr

Glycosylation: N linked vs O linked occur at which AAs?

N linked: Asn

O linked: Ser, Thr

Which proteins would get glycosylated?

Plasma membrane, secretory, lysosomal proteins

What are NDP-sugars?

nucleotide + 2 phosphates + sugar

Glycosyltransferases transfer _____ from NDP-sugar onto ______

Glycosyltransferases transfer sugar from NDP-sugar onto target protein

Where does glycosylation take place?

ER and Golgi

From the O type, which sugars get added to form the A and B blood type?

A: GalNAc

B: Gal

N linked glycosylation:

1. After the polypeptide is synthesized, it moves to the ____.

2. Here, a sugar oligosaccharide is added to ______.

N linked glycosylation:

1. After the polypeptide is synthesized, it moves to the ER lumen.

2. Here, a sugar oligosaccharide is added to Asn.

Trimming of N linked glycosylation

Some sugars are removed to help with protein folding

Golgi modifications of N linked glycosylation:

1. Sialic acid

2. Mannose 6 phosphate

1. Secretory and membrane proteins end with sialic acid

2. Lysosomal proteins get a mannose 6 phosphate, where the mannose is then cleaved, leaving the phosphate group

___ linked glycosylation occurs after ___ linked.

O after N

Glycosylation protects against _____

proteolysis

Mucin is an example of ___-linked oligosaccharides that shield epithelial cells in the airway from infection.

O

The viral protein on influenza attaches to _____ residues on glycoprotein. Neuraminidase ________ so the virus can keep moving around and infecting.

sialic acid

cleaves sialic acid residues after infection

Congenital diseases of glycosylation (CDG) are _____ disorders of ___ linked glycosylation.

autosomal recessive

N-linked

Myristoylation and palmitoylation are (irreversible/reversible).

What’s the purpose of these?

Myristoylation: irreversible

Palmitoylation: reversible

Increases hydrophobicity of proteins so they can bind the plasma membrane

Myristoyl-CoA

Donates a C14 Fa to a Gly residue

Palmitoyl-CoA

Donates a C16 FA to a Cys residue

Farnesyl pyrophosphate donates a _____ to a _____ consensus sequence of a target protein. In this process, ___ gets methylated.

donates a C15 hydrocarbon

CAAX sequence

C gets methylated

Hutchinson-Gilford Progeria is caused when the farnesyl lipid anchor is _______, leading to a ______ phenotype.

not cleaved

aging-like

T/F: cytosolic and ER ribosomes are identical. Explain

T - their location is determined by the nature of the protein that they translate

Cytosolic, ER, and secretory pathways

Cytosolic: proteins synthesized on free ribosomes

ER: proteins synthesized in ER that stay there

Secretory: proteins synthesized in ER that leave

Proteins have signal sequences that tell it _______.

where to go after translation

Mitochondrial DNA is encoded in the ______ and then transported after translation.

nucleus

Post translational transport of mitochondrial proteins into mitochondria:

1. ______ chaperone proteins keep the proteins unfolded

2. ____ complex recognizes it and allows entrance to matrix.

3. matrix _____ inside the matrix pulls the protein in.

4. Protease ____ the pre-sequence.

5. Protein folds (inside/outside) the matrix.

Post translational transport of mitochondrial proteins into mitochondria:

1. Hsp70 chaperone proteins keep the proteins unfolded

1. TOM complex recognizes it and allows entrance to matrix.

2. matrix Hsp70 inside the matrix pulls the protein in.

3. Protease cleaves the pre-sequence.

4. Protein folds inside the matrix.

Nuclear import of proteins goes through the ______ complex

nuclear pore complex

What is the protein sequence that tells it to be transported to the nucleus?

Nuclear localization signal (NLS)

Nuclear import of proteins:

1. _____, a receptor, binds NLS and transports it to the nucleus.

2. ____-GTP removes this receptor and imports it back to the ___.

3. The NLS (is/is not) removed after transport.

Nuclear import of proteins:

1. Importin, a receptor, binds NLS and transports it to the nucleus.

2. Ran-GTP removes this receptor and imports it back to the cytosol.

3. The NLS is not removed after transport.

T/F: both mitochondrial and nuclear import require energy (ATP)

T

Is the protein unfolded or folded when it gets imported to the mitochondria vs nucleus?

Mitochondria: unfolded

Nucleus: folded

In peroxisomal import, where does the receptor end up after bringing protein in?

Comes back out to the cytosol to be Ub and recycled

The cytosolic pathway involves transport to which 3 organelles?

Mitochondria, nucleus, peroxisome

Proteins in the secretory pathway have a _____ signal sequence that are recognized by ______.

N-terminal

signal recognition particle (SRP)

Proteins in the secretory pathway experience translocation. Explain.

Moves into ER lumen as its being translated

T/F: all protein translation starts in cytoplasm

T - some stay, some move to RER

How are proteins that are meant to be translated in the RER moved from cytosol → RER?

SRP recognizes their signal sequence and relocates it to the RER

What if a protein is being synthesized in the RER that’s meant to stay as a transmembrane protein?

The hydrophobic a-helix gets translated, the channel closes, and the rest of the chain is synthesized in the cytosol

Proteins are shipped from ER → golgi via ____ in the secretory pathway. This is done by the ______ of _____.

vesicles

the budding of vesicles

Proteins carrying a C-terminal KDEL

Remain in the ER (don’t go to golgi)

COPI vs COPII vesicles: direction, location

COPII: anterograde, ER → golgi (away from cell body)

COPI: retrograde, golgi → ER (towards cell body)

Clathrin-coated vesicles mediate transport from ___ → _____

golgi → lysosome

T/F vesicles stay coated as they transport proteins from ER/golgi

F - become uncoated after budding

v-SNAREs and t-SNAREs

vSNAREs are on the surface of vesicles and recognize tSNAREs of the target location

lock and key

From the golgi, what is the constitutive vs regulated secretory pathway

Constitutive: default pathway to cell surface

Regulated: cargo is stored in secretory granules and released when signaled to

Tetanus and botulinum toxins cleave ____, causing no vesicle function.

SNARE

Tetanus toxin blocks release of _____ from ___ neurons, resulting in spastic paralysis.

GABA, inhibitory

Botulinum toxin blocks release of ___ from _____ neurons, resulting in flaccid paralysis.

ACh, excitatory

Proteins need a ____ tag to be sent to lysosomes.

M6P

Clathrin can help with the ____ of molecules and merge with early endosomes.

endocytosis