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Structure/Function of the

Organelles - Synthesis

Protein Synthesis and Transport within the Cell


The Endomembrane System
Membrane organelles a dynamic, integrated network
Materials are shuttled small transport vesicles
Bud from a donor membrane compartment Move in a directed manner Pulled by motor proteins Tracks microtubules of the cytoskeleton At their destination fuse with membrane of acceptor compartment

Repeated cycles of budding and fusion

Protein Synthesis and Transport within the Cell


The Endomembrane System

Protein Synthesis and Transport within the Cell


The Endomembrane System
Typical biosynthetic pathway
Proteins synthesised in the endoplasmic reticulum Modified through the Golgi complex Transported to destination

Also a secretory pathway

Protein Synthesis and Transport within the Cell


The Endomembrane System
Secretory activities two types
Constituitive
Transported from site of synthesis and secreted continuously

Regulated
Materials to be secreted are stored in large, densely packed, membrane-bound secretory granules Discharged following specific stimulus Examples: endocrine cells, acinar cells, nerve cells

Protein Synthesis and Transport within the Cell


The Endomembrane System
Materials moved out by secretory pathways Materials moved in by endocyte pathway
From outside the cells to endosomes / lysosomes

Secreted Proteins targeted by sorting signals


In the aa sequence or Attached oligosaccharide

Protein Synthesis and Transport within the Cell

The Endomembrane System


Autoradiography
Example: Acinar cells synthesis of digestive enzymes

Protein Synthesis and Transport within the Cell The Endomembrane System
Autoradiography
Cells overlayed with photographic film Exposed to radioisotopes in the sample Radiolabeled aa

Protein Synthesis and Transport within the Cell The Endomembrane System
Autoradiography
Tissues washed and transferred to unlabelled aa pulse-chase

Protein Synthesis and Transport within the Cell The Endomembrane System
Biochemical analysis of subcellular fractions
Technique to homogenize cells
Cytoplasmic membranes fragmented and form vesicles Types separated using cell fractionation ER and Golgi form microsomes Fractioned into rough and smooth

Protein Synthesis and Transport within the Cell


The Endomembrane System
The Endoplasmic Reticulum (ER)
Two categories
Rough (RER) Smooth (SER)

Composition of luminal or cisternal space unique RER ribosomes bound to cytosolic surface SER no ribosomes RER & SER many complex differences

Protein Synthesis and Transport within the Cell


The Endomembrane System
RER vs SER
RER
Extensive Interconnected sacs cisternae Continuous with nuclear envelope

SER
Tubular Interconnecting pipelines

Protein Synthesis and Transport within the Cell


The Endomembrane System
SER functions
Skeletal muscle Kidney tubules Steriod producing cells Liver
Detoxification Glucose-6-phosphatase

Protein Synthesis and Transport within the Cell


The Endomembrane System
RER functions Protein Synthesis Polypeptides synthesized at two locales
1. On ribosomes attached to cytosolic surface of RER
Proteins secreted from the cell Integral membrane proteins Soluble proteins within ER, Golgi, lysosomes, endosomes

Protein Synthesis and Transport within the Cell


The Endomembrane System
RER functions Protein Synthesis Polypeptides synthesized at two locales
2. Other polypeptides on free ribosomes
Released into the cytosol Enzymes Peripheral proteins plasma membranes Proteins nucleus Protein mitochondria, peroxisomes

Protein Synthesis and Transport within the Cell


RER functions Protein Synthesis
How are proteins synthesized at different sites?
Secretory proteins signal sequence Polypeptide moves to cisternal space of ER
cotranslationally

Protein Synthesis and Transport within the Cell RER functions Protein Synthesis
Begins after a messenger RNA binds to a free ribosome

Protein Synthesis and Transport within the Cell RER functions Protein Synthesis
Other points to note
Interaction between signal sequence and SRP a key step SRP is the tag enables specific binding of the complex to the ER Binding through at least two interactions
SRP SRP receptor Ribosome and translocon

Following binding
Nascent peptide released from SRP Inserted into translocon

On termination
Ribsome released Translocon channel reverts to narrow conformation

Protein Synthesis and Transport within the Cell RER functions Protein Synthesis

Protein Synthesis and Transport within the Cell RER functions Processing of proteins
Polypeptide is moved from the translocon to the cisterna
Enzymes in the membrane
Signal peptide removed signal peptidase Carbohydrates added oligosaccharyltransferase

Protein Synthesis and Transport within the Cell RER functions Processing of proteins
In the lumen
Molecular chaperones BiP (binding protein) Recognize and bind unfolded proteins Aid in attaining native conformation Movement of the protein into the ER lumen Protein disulfide isomerase (PDI)

Protein Synthesis and Transport within the Cell RER functions Processing of proteins
Misfolded proteins
Transported to cytosol Destroyed in proteasomes
Quality control

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