5. Endoplasmatic reticulum and protein synthesis Flashcards

1
Q

The endoplasmatic reticulum

A
  • A network of membranous tubules within the cytoplasm, which is continuous with the nuclear membrane.
  • First observed in 1945 (Porter, Claude & Fullam

Percentage of total cell volume: 12
Approximate number per cell: 1

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2
Q

Endoplasmatic reticulum

A
• Involved in protein and lipid synthesis
• Present in most eukaryotic cells
• Two types of ER
– Rough ER
– Smooth ER
  • RER is studded with ribosomes and is involved in protein synthesis
  • SER does not have ribosomes and mostly deals with lipid synthesis
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3
Q

Structure of the ER

A
  • Membranous network of cisternae
  • Phospholipid membrane encloses the cisternal space (lumen), which is continuous with the perinuclear space, but separate from the cytosol
  • Relative quantities of RER and SER can change from one cell type to another, depending on changing metabolic activities
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4
Q

Protein synthesis

A

• Involves the translation of message (mRNA) into protein
– Performed by a ribosome

• Ribosome binds to the mRNA that has exited the nucleus

• tRNA, which have amino acids attached, work with the ribosome to add amino acids one at a time to build the new (nascent) protein
– There are 20 tRNA, one for each amino acid

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5
Q

The ribosome

A

• Molecular machine found in all living cells

• Primary function is protein synthesis
– Ribosomes link amino acids together to form proteins

• Composed of 2 major subunits (40S & 60S in eukaryotes)
– Small subunit reads the mRNA
– Large subunit adds amino acids to the growing polypeptide chain (catalysed by the catalytic peptidyl transferase activity of the subunit)

• Each subunit composed of one or more rRNA and a variety of proteins
– Collectively termed the translational apparatus

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6
Q

Translation specifics

A

-

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7
Q

The tRNA + aminoacyl tRNA synthetase

A

-

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8
Q

Polyribosomes

A
• Clusters of ribosomes that bind the same mRNA molecule
• Can be found in three forms
– Free
– Membrane-bound
– Cytoskeletal-bound
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9
Q

The ER + protein trafficking

A

• Secretory pathway – involves the ER
– Soluble and membrane proteins

  • Proteins secreted from the cell
  • Proteins resident in the lumen of the ER and Golgi complex
  • Integral proteins in the membranes of the ER, Golgi & plasma membrane

• Non-secretory pathway
– Targeting to an intracellular organelle (mitochondria, nucleus, etc)

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10
Q

The secretory pathway (involves ER)

A

-

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11
Q

Overview of secretory pathway

A

• Ribosomes synthesising proteins with an ER signal sequence are directed to the ER
– Facilitated by the signal recognition particle (SRP)

• After translation and translocation are complete in the ER, proteins are packaged into transport vesicles destined for the Golgi
– Further sorted for eventual delivery to the plasma membrane or lysosomes

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12
Q

Signal sequences

A

Import into ER:

  • > +H3N - indication of N-terminus of protein
  • > extended block of hydrophobic amino acid (e.g. Leu)
  • > negatively charged (e.g. Glu)

Retention in lumen of ER:

  • > positively charged (e.g. Lys)
  • > negatively charged (e.g. Asp)
  • > COO- - indication of C-terminus

Import into mitochondria:

  • > +H3N - indication of N-terminus of protein
  • > positively charged (e.g. Arg)

Import into nucleus:
-> positively charged (e.g. Lys)

Import into peroxisomes:
-> positively charged (e.g. Lys)

=> The ER retention signal commonly referred to by single-letter amino acid abbreviation, KDEL.

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13
Q

ER - targeted protein synthesis

A
  • > mRNA encoding cytosolic protein remains free in cytosol
  • > elongating polypeptide chain
  • > polyribosome free in cytosol
                                  ¦

common pool of ribosomal subunits in cytosol

                                ¦

ER membrane with the ER lumen
-> ER signal sequence
-> mRNA encoding a protein targeted to ER remains membrane-bound
=> polyribosome bound to ER membrane by multiple nascent polypeptide chains

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14
Q

Signal recognition particle function

A

mRNA signal sequence on nascent polypeptide + tRNA -> binding of SRP to signal peptide causes a pause in translation -> SRP bound ribosome attaches to SRP receptor in ER membrane

  • SRP recognises signal sequence on nascent polypeptide
  • SRP binding to signal sequence stalls translation whist complex moves to ER membrane and binds to SRP receptor
  • SRP/Ribosome complex associates with SRP receptor on RE
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15
Q

SRP function

A

=> translation continues and translocation begins
=> SRP and SRP receptor displaced and recycled

  • SRP docks with SRP receptor
  • SRP transfers nascent peptide and ribosome to the translocon (Protein Translocator)
  • SRP and SRP receptor dissociate and are recycled
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16
Q

The signal recognition particle

A
  • > translational pause domain
  • > SRP RNA molecule
  • > hinge
  • > signal sequence binding pocket with signal sequence
17
Q

Fate of the ER signal sequence

A

-> NH2

Inactive protein -> active translocator -> signal peptidase

  • > NH2 in cytosol
  • > NH2 / COOH in ER lumen (mature soluble protein in ER lumen)
18
Q

Further modification

A

• Protein folding in the ER
– Formation of S-S bonds
– Chaperones (BiP, calnexin, calreticulin)
– Peptidyl prolyl isomerases

• Oligosaccharide decoration and processing -PTM
– Most proteins in ER are glycosylated
– N-linked glycosylation