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Frontiers in infectious diseases > 15. Bacterial protein secretion and assembly of surface structures > Flashcards

15. Bacterial protein secretion and assembly of surface structures Flashcards

1
Q

What are most bacterial protein secretion systems important in?

A

Pathogenesis

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2
Q

Why is secretion/transport of molecules so important for bacteria?

A
  1. To cause disease bacteria must interact with their host and they use molecules to do this.
  2. They sense their host’s physiological cues.
  3. They need to deliver bioactive molecules (toxins or effector) to host cell/tissues.
  4. They do this by making a structure that secretes or delivers molecules to host tissues/cells.
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3
Q

How are bacterial effector molecules transported from the cytoplasm?

A
  1. They are transported through the membrane.
  2. This is more complicated in gram negative bacteria due to its membrane complexity.
  3. Some proteins integrate into the membranes and these have important roles in pathogenesis and toxicity.
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4
Q

What proteins are integrated into the membrane but are not integral to it?

A
  1. Proteins that have functions important for pathogenesis.
  2. An example of these are secretion systems.
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5
Q

What secretion systems do gram positive bacteria use?

A
  1. Sec
  2. Tat
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6
Q

What do gram-negative bacteria secretion systems have to do?

A
  1. As their membranes are more complex so are their secretion systems.
  2. The 2 membranes have to be linked to transport proteins through both.
  3. This needs to be synchronised.
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7
Q

What secretion systems do gram-negative bacteria use?

A
  1. They use Sec and Tat linked to other secretion systems.
  2. They use 2 step secretion systems like type 5.
  3. They use 1-step secretion systems like type 3.
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8
Q

What are 2-step gram negative secretion systems?

A
  1. They use Sec and Tat to transport things across the inner membrane.
  2. They then use secretion systems to transport proteins across the outer membrane.
  3. Examples are type 1, 2, and 5 which secrete proteins like proteases.
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9
Q

What are 1-step gram negative secretion systems?

A
  1. They transport proteins from the bacterial cytoplasm directly to the host cell cytoplasm.
  2. They are big structures made up of lots of proteins.
  3. The effect molecules have no contact with the external space.
  4. Examples are type 3, 4 and 6 secretion systems.
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10
Q

How does protein secretion work in Gram-Positive Bacteria?

A
  1. Secretion systems used by pathogenic bacteria are essential for their virulence.
  2. These secrete toxins and adhesins and other virulence factors.
  3. Use 3 main pathways.
  4. The general secretory pathway or Sec.
  5. The Twin-arginine translocation pathway or Tat.
  6. The ATP-binding cassette transporter
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11
Q

Where is the Sec pathway found?

A
  1. It is ubiquitous and essential for life.
  2. It is used by both gram-positive and gram-negative bacteria.
  3. It is found in all 3 domains of life: bacteria, archaea and
    eukaryotes.
  4. It is also found in eukaryotic organelles for transport across organelle membranes.
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12
Q

What does Sec-dependent secretion require?

A
  1. Proteins require a signal/leader peptide sequence to be secreted.
  2. This guides them and allows them to be translocated across the cytoplasmic membrane.
  3. The proteins have to be unfolded.
  4. Transport requires hydrolysis of ATP.
  5. The signal peptide needs to be removed once it is secreted.
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13
Q

What is the mechanism of the Sec pathway?

A
  1. The pre-protein with the signal sequence is targeted to the cytoplasmic membrane.
  2. This is aided by the export chaperone SecB.
  3. SecB leads the pre-protein to SecA on the inside of the membrane.
  4. SecA is an ATPase.
  5. Every time an ATP is hydrolysed the pre-protein is pushed ~20 amino acids into the SecYEG channel.
  6. Once the pre-protein has been translocated across the membrane, the signal peptide is cleaved.
  7. This is done by the type 1 signal peptidase, possibly SecDF.
  8. Then the bacteria is folded by other secreted proteins.
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14
Q

Why is the chaperone needed in the Sec pathway?

A

Due to pre-proteins being unstable as they are not folded.

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15
Q

Where is the Tat pathway found?

A
  1. It is not as widespread as Sec.
  2. It is in many bacterial and archaeal organisms.
  3. It is also in plant cell thylakoid membranes (chloroplasts).
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16
Q

What proteins does Sec transport?

A

Unfolded proteins

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17
Q

What proteins does Tat transport?

A

Folded proteins

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18
Q

What drives Sec?

A

ATP hydrolysis

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19
Q

What drives Tat?

A

The proton motive force.

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20
Q

What does the Tat secretion system require?

A
  1. Folded proteins
  2. Proton motif force
  3. A signal peptide
  4. A signal motif within the protein
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21
Q

What signals guide peptides to the Tat pathway?

A
  1. There is the 40 amino acids signal peptide.
  2. The proteins also contain a motif specific for Tat transport.
  3. This sequence is serine or theronine/arginine-arginine/x/phenylalanine/leucine/lysine.
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22
Q

What is the mechanism of the Tat pathway?

A
  1. Before export, Tat exported proteins are folded into their conformation.
  2. They contain N-terminal signal sequence with the twin arginine motif.
  3. The folded pre-protein is recognised by the TatB and TatC complex.
  4. This complex delivers the pre-protein to TatA.
  5. TatA is the channel in the membrane that translocates the proteins across the membrane.
  6. After export, the signal sequence is removed by signal peptidase.
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23
Q

What is the difference between SecYEG and TatA?

A

TatA is a bigger channel than SecYEG as it transports folded proteins, which are bulkier than unfolded proteins.

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24
Q

What are the similarities between Sec and Tat?

A
  1. Both are highly conserved
  2. Both use pre-proteins with signal peptides.
  3. Signal peptides are cleaved on export.
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25
What are the differences between Sec and Tat?
1. Sec transports unfolded proteins using ATP hydrolysis. 2. Tat transports folded proteins using the proton motive force.
26
What are the different secretion systems in gram-negative bacteria?
1. Tat 2. Sec 3. Type 1 - 9 secretion systems.
27
Gram-negative secretion systems: Tat-pathway
Used for export of fully folded proteins with metal co-factors bound
28
What is the only pathway that transports folded proteins?
Tat
29
Gram-negative secretion systems: Sec
Used to transport proteins into the periplasm and across the inner membrane.
30
Gram-negative secretion systems: Type 1 secretion systems
ABC-transportors that are used for haemolysin secretion.
31
Gram-negative secretion systems: Type 2 secretion systems
1. Main terminal branch of Sec for transport across the outer membrane. 2. Used for cholera toxin secretion. 3. Used for type 4 pili biogenesis.
32
Gram-negative secretion systems: Type 3 secretion systems
Used as flagella and virulence.
33
Gram-negative secretion systems: Type 4 secretion systems
1. Used for DNA conjugation 2. Pertussis toxin secretion and virulence
34
Gram-negative secretion systems: Type 5 secretion systems
1. Autotransporters. 2. Outer membrane insertion 3. Secretion of virulence proteins
35
Gram-negative secretion systems: Type 6 secretion systems
1. Phage derived 2. Contact dependent killing in bacteria. 3. Virulence
36
Gram-negative secretion systems: Type 8 secretion systems
1. For small curli like cilia 2. Biofilm formation
37
Gram-negative secretion systems: Type 9 secretion systems
1. Newly discovered 2. Secrete adhesive filaments like motility 3. Secrete peptidases called gingipains
38
What are the 2 main mechanisms of getting molecules out of the gram-negative bacteria cytoplasm?
1. Sec-dependent transport or periplasmic intermediate. 2. Sec-independent transport or no periplasmic intermediate
39
What are the 2 steps in Sec-dependent gram negative secretion?
1. Translocation across the inner membrane using Sec or Tat. 2. Translation across the outer membrane using a secretion system.
40
Examples of Sec-dependent secretion systems
1. T2SS 2. T5SS 3. T4 pilus 4. T7SS 5. T8SS
41
What is the process of Sec independent gram-negative transport?
1. There is 1 step from the bacterial cytoplasm to the extracellular space. 2. The Nanomachine spans both membranes and the periplasm. Examples: T1SS, T3SS, T4SS, T6SS, T9SS.
42
What are the 3 main classes of Type 5 secretion systems?
1. Autotransporter 2. Two-partner secretion 3. Chaperone-usher (not used in pathogenesis).
43
What do proteins secreted through T5SSs have?
1. An N-terminal Sec or Tat signal sequence as it is a Sec dependent secretion system. 2. This is cleaved off as they pass into the periplasm.
44
What is the mechanism of the T5SS autotransporter?
1. T5SS autotransporters contain all the information needed to transport themselves across the membrane in their sequence. 2. This includes the T5SS. 3. They have a signal sequence that targets them to Sec. 4. The N terminal passenger domain is the functional domain. 5. The C terminal translocator domain forms a ß barrel. 6. The protein uses Sec to cross the inner membrane. 7. The translocator forms a channel in the outer membrane and the passenger domain passes through the channel to exit the bacteria. 8. Once the passenger passes through, it can be cleaved or remain attached.
45
What are some examples of T5SS autotransporters?
1. Adhesins 2. Degradative enzymes 3. Cytotoxins 4. N. gonorrhoeae IgA protease.
46
What is the T5SS two-partner system?
1. This uses 2 proteins: 1 partner contain the domain and makes the ß-barrel, 1 partner is secreted. 2. This system uses Sec to cross the inner membrane 3. TpsB protein family form the translocator in the outer membrane. 4. TpsA secreted protein family is the passenger protein that is secreted.
47
What are TpsB proteins?
1. A protein in T5SS two-partner system. 2. It is a channel forming ß barrel. 3. 500-800 amino acids long 4. Highly conserved proteins.
48
What are TpsA proteins?
1. Large ~3000 amino acids long 2. The signal peptide is cleaved in the periplasm. 3. Tps domain at the N terminus 4. Get released into the extracellular space. 5. Cross the outer membrane through TpsB. 6. Repeated sequences that form helical structures.
49
What is the T3SS similar to?
1. The flagella 2. This similarities is in structure not function.
50
What induces secretion in T3SS?
Host-cell contact
51
What is the T3SS?
1. A complex nanoinjector. 2. It is made up of more then 20 proteins. 3. It is involved in pathogenesis by secreting effector molecules into host cells. 4. The complexity means the genes encoding it are close together on a pathogenicity island. 5. Can be plasmid or chromosomal 6. Evolutionary relationship with flagella.
52
What is the mechanism of T3SS?
1. 1 set of protein rings in the inner membrane. 2. 1 set of protein rings in the outer membrane. 3. They are connected by protein repeats in the periplasm. 4. The tip of the middle connector are the translocon proteins. 5. The translocon inserts and integrates into the host membrane. 6. The syringe part is hollow so proteins can be transported through the middle.
53
What is the function of T3SS?
To inject virulence proteins into a target cell.
54
What drives T3SS translocation?
1. ATP hydrolysis and the proton motive force. 2. This drives the rotation of the proteins and the movement of proteins.
55
How many secretion system can a bacteria have?
1. 1 or lots. 2. They can have lots of the same type in their membrane. 3. They can also have lots of different types expressed. 4. This is due to the fact they have different functions.
56
What waste do secretion systems that integrate into host membranes generate?
1. They are generally only activated on contact with or sensing of the host cell. 2. It is a directed response to a cell. 3. The systems are pre-formed but only activated when needed. 4. As this is direct no proteins are waste into the extracellular space.
57
How does E. coli use the T3SS?
1. It uses T3SS to transport effector proteins like Tir to cause changes in the host cell and these are transported through the translocon. 2. Once they enter the host cell the proteins fold and take their conformation 3. Tir integrates into the host cell membrane. 4. Tir binds to intamin on the bacteria membrane. 5. When Tir binds intamin it causes changes in the host cell actin. 6. This pushes the host cell membrane out to form a pedestal. 7. This enhances contact with the bacteria and facilitates entry.
58
What does shigella use T3SS for?
To induce membrane ruffles into host cells for entry.
59
What is the type 6 secretion system?
1. It is pre-formed in the bacteria 2. On contact they sense a threat and the translocon is pushed to the host cell membrane. 3. This delivers toxins to other cells. 4. These can be bacteria or eukaryotes. 5. Once this has happened, the T6SS is disassembled.
60
What are T6SS mainly used for?
Used by bacteria to eliminate competition in the environment.
61
What can bacteria attack with a T6SS cause?
1. It can cause retaliation from bacteria also with T6SS. 2. The more virulent bacteria will survive.
62
What else can T3SS be used for?
1. It could be used as a tool for therapeutic drug delivery 2. It can be used to target specific cells. 3. It has 1 step secretion so the drug is delivered directly.
63
How could T3SS be used in cancer immunotherapy?
1. Cancer cell grow in low oxygen environment which is good for bacteria. 2. A protein produce and expressed by cancer can be expressed in a bacteria. 3. A T3SS could be used to transport this cancer protein into a APC. 4. This cancer antigen is then processed and presented on the APC to prime the immune response to target cancer cells. 5. This looks for a targets cancer antigens using CD8+ T cells. 6. This is generally safe for human use and worked in clinical assays.
64
How could T3SS be used in cellular reprogramming?
1. You can change the gene expression in a specific cell. 2. You can use a T3SS to deliver transcription factors to alter the gene expression 3. This can be used to make pluripotent stem cells.
65
How could T3SS be used in passive immunotherapy?
1. Get the bacteria to make antibodies against the pathogenic proteins on a disease cell. 2. Use a T3SS to inject antibodies into infected cells. 3. This is targeted treatment.
66
How could T6SS be used therapeutically?
1. It is activated only when they sense threats. 2. If you can program them to sense other threats like diseased cells you can use them to make targeted therapies. 3. Can be used to inject antifungals in to candida cells/infection. 4. Can also be used to enter the vacuole of macrophages and inject molecules that alter actin cytoskeleton in the macrophage.
67
What is bacterial nanowire?
1. Shewanella oneidensis can make protrusions with the potential to generate electricity. 2. These are still attached to the bacteria. 3. Normally the electrodes don’t touch so no electric circuit is made. 4. When they do touch it can be used to generate electricity. 5. This can have potential applications.

Frontiers in infectious diseases (19 decks)

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