MCB 2050 Endomembrane System L8-L9 Questions With Complete Solutions

MCB 2050 Endomembrane System L8-L9 Questions With
Complete Solutions

COPII-coated vesicle assembly at the EREs

What are steps 3 and 4? Correct Answers Step 3:
- after release of nascent COPII vesicle from 'donor' membrane
(i.e., ER/ERES), Sec23 promotes hydrolysis of GTP by Sar1
(GTPase)
- Sar1-GTP converted to Sar1-GDP

Step 4:
- GTP-hydrolysis by Sar1 results in disassembly of COPII
protein coat
--> Sar1-GDP and all other COPII proteins released into
cytoplasm for additional rounds of COPII-coat assembly at
ERES
- results in nascent, uncoated vesicle...

COPII-coated vesicle assembly at the EREs

What is step 1?

What is Sec12? Correct Answers - soluble COPII component
Sar1 (GTPase - binds GDP and GTP) is recruited from the
cytoplasm to ER (ERES) membrane via its binding to Sec12
--> Sec12 is ER integral membrane protein - functions as
guanosine-exchange factor (GEF) that catalyzes exchange of
GDP for GTP on Sar1

,- Sar1 binding to GTP (Sar1-GTP) causes conformational
change - exposes Sar1 amphipathic hydrophobic N-terminus
(serves as ER membrane 'anchor')
- Sar1-GTP integrated into outer (cytoplasmic-facing) leaflet of
ER membrane bilayer

COPII-coated vesicle assembly at the EREs

What is step 2?

What is the role of Sec24? Correct Answers - Sar1-GTP binds
(recruits) several other COPII proteins from cytosol to ER
membrane surface
- Sar1 initially recruits Sec23 and Sec24
--> soluble proteins form a ternary complex with Sar1 at ER
membrane surface
--> act as a structural 'scaffolding' and begin to promote outward
(i.e., towards cytosol) bending of ER membrane
--> beginning of COPII vesicle 'bud' formation

- Sec24 also involved in vesicle 'cargo' protein selection
- Sec24 binds to cytoplasmic-facing domains of various selected
ER integral (trans) membrane proteins:
--> membrane 'cargo' proteins - destined to exit ER for Golgi
--> membrane 'cargo-receptor' proteins - bind (via lumenal-
facing domains) to soluble (lumenal) 'cargo' proteins destined to
exit ER for the Golgi
--> membrane 'trafficking' proteins (e.g., v-SNAREs) - required
for subsequent trafficking and docking of nascent vesicle with
proper 'acceptor' membrane (i.e., Golgi)

, - selection (recognition) of vesicle membrane cargo by Sec24
mediated by an ER export sorting signal
--> most common is di-acidic ER export signal (-Asp-X-Glu-)
- located in cytoplasmic-facing domains of Sec24- selected
membrane proteins
- ER export sorting signals not found on ER resident proteins
--> various other sorting signals (sequences) responsible for
proper localization of proteins in endomembrane system
- all Sec24-bound proteins (and soluble proteins bound by
membrane cargo-receptor proteins ) concentrated within
growing, COPII protein-coated vesicle 'bud'

Core modification

What happens during N-linked glycosylation and modification?
Correct Answers - during N-linked glycosylation and
modification, the nascent protein is rapidly folded into the
proper 3D conformation
- mediated by several ER lumen and membrane proteins
- reticuloplasmins - ER molecular chaperones, including BiP,
calreticulin and calnexin
--> bind transiently (reversibly) to nascent ER proteins to
prevent misfolding or aggregation
- protein disulfide isomerase (PDI) - enzyme catalyzes the
formation of intra/intermolecular disulfide bonds
--> disulfide bonds between cysteine residues on the same or
different (nascent) polypeptides promote proper folding and
assembly by stabilizing their proper 3D conformation
- core oligosaccharide(s) added to nascent protein during N-
linked glycosylation also contribute to proper protein