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BIS 104 MIDTERM 2 > Lecture 9 > Flashcards

Lecture 9 Flashcards

1
Q

What is the signal sequence hypothesis?

A

Microsomes added to proteins normally secreted out of the cell –> protein is smaller b/c signal peptide gets cleaved –> lower on SDS PAGE gel than w/o microsomes

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2
Q

If a protein is higher on the SDS PAGE, what can be said about its signal peptide? Lower?

A
  • Higher = protein has a signal peptide

- Lower = signal peptide has been processed/cleaved

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3
Q

How can you identify on the SDS PAGE a protein that is normally secreted?

A

Normally secreted proteins treated w/ microsomes and protease have a band the same height as when treated w/ just microsomes

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4
Q

List the 6 major players in the mechanism of protein translocation.

A
  • mRNA
  • Ribosome
  • Signal peptide
  • SRP
  • SRP receptor
  • Pore in the ER
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5
Q

What is the role of mRNA in protein translocation?

A

Codes for the protein to be translocated across ER membrane

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6
Q

What is the role of ribosomes in protein translocation?

A

Translates mRNA into a peptide

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7
Q

What is the role of the signal peptide in protein translocation?

A

Targets the protein/mRNA/ribosome complex to the ER or translation and translocation

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8
Q

What is a signal peptide composed of?

A

6-15 hydrophobic residues

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9
Q

Where are signal sequences normally found on a secreted protein? Transmembrane protein?

A
  • Normally @ the N-term of a secreted protein

- Can be found in the middle of a transmembrane protein

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10
Q

What does the location of a signal sequence on a protein dictate?

A

Topology of the protein

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11
Q

What is SRP? Role in protein translocation?

A

Signal Recognition Particle: recognizes signal sequence, arrests translation, and targets ribosome to ER membrane

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12
Q

What is SRP composed of?

A

6 peptides and 1 RNA molecule

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13
Q

What is a major component of the ER pore?

A

Sec61

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14
Q

Where does all protein synthesis occur? Why?

A

All protein synthesis occurs in the cytoplasm b/c all ribosomes are in the cytoplasm

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15
Q

Describe the process of protein translocation across the ER membrane?

A
  1. SRP in cytoplasm (GDP state) binds to signal sequence on mRNA/ribosome/protein complex
  2. Ribosomes act as a GEF for SRP –> converts from GDP to GTP state
  3. SRP acts as GEF2 for SRP receptor –> converts from GDP to GTP state
  4. Complex binds to SRP receptor b/c both in GTP state
  5. Translocon (Sec61) functions as GAP –> both SRP and SR convert from GTP to GDP
  6. SR releases complex to translocon
  7. SRP is released from complex
  8. Plug on translocon opens –> allows unfolded peptide to enter ER lumen
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16
Q

You have isolated a yeast strain with a mutation in the GTP binding domain of SRP. This mutant SRP can not bind a nucleotide. Where would you predict to find a protein that is normally secreted?

A

In the cytoplasm

17
Q

If you add nuclear localization sequence PKKKRKV to the middle of BiP protein that resides in ER lumen, where would you expect to find BiP?

A

ER b/c NLS overriden by signal sequence for ER since receptors will read what it on the end first

18
Q

How does the pore of the translocon contribute to topology?

A

CHARGES: pore surrounded by charged residues on cytoplasmic surface –> protein w/ opposite charged residues will end up on the cytoplasmic side of the membrane

19
Q

What are the characteristics of Type I membrane proteins?

A
  • N-term in ER lumen
  • C-term in cytosol
  • Has signal sequence
  • Has stop transfer sequence (STS) in the middle
  • Single pass
20
Q

What are the characteristics of Type II membrane proteins?

A
  • N-term in cytosol
  • C-term in ER lumen
  • Internal start signal sequence (ISS)
  • (+) charged AAs before ISS
  • Single pass
21
Q

Which topology of membrane proteins is relatively rare?

A

Type III

22
Q

What are the characteristics of Type III membrane proteins?

A
  • N-term in ER lumen
  • C-term in cytosol
  • No signal peptide
  • (+) charged AAs after ISS in cytosol
  • Single pass
23
Q

Which 2 membrane protein topologies are similar?

A

Type I and III

24
Q

What is the difference b/t Type I and III membrane protein topologies?

A
  • Type I use N-term signal sequence to co-translationally translocate N-term into ER lumen
  • Type III use internal signal w/ (+) charges to determine its topology and pushes N-term into the ER after translation
25
Q

What are the characteristics of all Type IV membrane proteins?

A
  • Multi pass
  • May or may not have signal peptide
  • Domains alternate b/t ER lumen and cytoplasm
  • C-term in cytoplasm or lumen based on # of membrane passes/spans
26
Q

What are the characteristics of Type IV-A membrane proteins?

A
  • N-term in cytosol

- ISS-STS-ISS-STS

27
Q

What are the characteristics of Type IV-B membrane proteins?

A
  • N-term in ER lumen
  • (+) charged AAs after 1st ISS
  • ISS-ISS-STS-ISS-STS or SSS-STS-ISS-STS-ISS
28
Q

What identifies the signal sequence on a hydrophobicity plot?

A

First peak

BIS 104 MIDTERM 2 (4 decks)

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