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Protein Sorting
ISAT 351, Spring 2004
College of Integrated Science and Technology
James Madison University
Intracellular Compartments and Protein Sorting
Many chemical reactions in the cell are mutually incompatible (protein synthesis and degradation)
How does the cell control these reactions? Intracellular compartments are used to segregate
and isolate different chemical reactionsHow do proteins know the correct
compartment and how are they transferred? Signal sequences direct protein traffic
Membrane -Bound Compartments
Endoplasmic reticulum (ER): synthesis and modification of lipids and proteins for distribution
Golgi apparatus: modification, sorting, and packaging of proteins for delivery
Lysosomes: intracellular degradationEndosomes: sorting of endocytosed materialPeroxisomes: oxidation of toxic molecules
Protein Sorting is in One Direction
Why is this so?•Amino acid sequence defines protein fate•Some proteins synthesized in cytosol , then transported; other proteins complete synthesis at organelle•Post-translational modification of protein
•Gradient of immature-to-mature protein may be localized in compartments
Protein Transport
Mechanisms
1. Transport through pores (nucleus)
2. Transport across membranes (chloroplast and mitochondria)
3. Transport by vesicles (ER and Golgi)
Protein Sorting Signal Sequences
Signal sequences are a continuous stretch of amino acids (15 to 60) within the protein to be sorted.
Specific sequences direct the protein to the Nu, MT, CP, peroxisomes, or ER
Cytosolic proteins lack the signal sequence
Nuclear Protein Transport
Nu proteins are synthesized in the cytosol and actively transported via Nu pores
Nuclear localization signal (+ charged sequence) unique to Nu proteins
Mitochondria Protein Transport
Nu-encoded proteins synthesized in cytosol and imported by Mt receptor
Protein unfolds during transport refolds internally
Signal sequence removed Similar mechanism for CP
Transport into the ER
Proteins enter the ER during protein synthesis ER lumen, ultimately for secretion ER membrane, ultimately for membrane
proteins
The ER signal sequence directs the ribosome to the RER
RER
Secretory Proteins are Synthesized across RER-M into the RER Lumen
Integration of Transmembrane Protein into Membrane
Post-translational Modification of Proteins in the RER
• Post-translational modifications of protein• Gradient of immature-to-mature protein may be localized
in compartments• Traffic is unidirectional, from ER to golgi
• In ER, protein is synthesized and modified• In golgi, protein is modified and sorted• Vesicle traffic (fission and fusion events) move protein,
ultimately to plasma membrane
ER Protein Glycosylation
Oligosaccharide side chains (sugars) are added to many proteins in the ER, producing glycoproteins
Functions of glycosylation: Protection from degradation Transport and packaging signals, Cell communication when displayed on the
outer membrane as glycocalyx
ER Glycosylation: Oligosaccharide Attachment
ER Glycosylation
Oligosaccharide may be further modified downstream
Transport vesicles carry glycoprotein to to golgi QC failures:
Cystic fibrosis: membrane protein improperly folded Alzheimer’s disease: improper clipping of amyloid
Transport Vesicles
Transport vesicles shuttle proteins between various organelles and to the plasma membrane (exocytosis)
Vesicles that bud from membranes have a distinct protein coat (coated vesicles) Specific marker proteins on the surface of vesicles
(SNAREs) bind to target membranes
Vesicles fuse to the target membranes and release the transported molecules
Vesicle Traffic
Golgi Apparatus
Golgi Apparatus Organization & Functions
• Stacks closest to ER (“cis” face) receive vesicles’ contents from ER
• Proteins modified (e.g., glycosylation or clipping) in subsequent cisternae
• Transport via series of fission and fusion events• Furthermost stacks (“trans” face) release vesicles that
travel to PM• Each compartment contains unique enzymes; thus,
gradient of immature to mature proteins
Transport Vesicle Docking is Mediated by Proteins
Transport Vesicle Fusion is Mediated by Proteins
Exocytosis Releases Secretory Proteins
Constitutive vs. Regulated Secretion
All cells are capable of constitutive secretion
Regulated secretion requires an extracellular stimulus Example: insulin
release
Endocytosis
Endocytosis: cells take up fluid, molecules, and other cells
Pinocytosis involves the ingestion of fluids, molecules, and small particles
Phagocytosis involves the ingestion of large particles and microorganisms
Ingested material is delivered to the lysosome
Phagocytosis
Specialized phagocytic cells (e.g.,
macrophages) can ingest invading microorganisms
Lysosome
•Lysomes contain hydrolytic enzymes that digest both intra-and extracellular materials•Enzymes are most active in acidic conditions•Not just a dump: Membrane recycling
Pathways to the Lysosome
Questions to Think About
How does compartmentalization contribute to protein sorting?
What are some consequences of misprocessing? What roles do proteins play in secretion?
Signals? Vesicle traffic?
How do membrane lipids recycle? (hint: endocytosis)
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