L4 - Microbial Transport and Communication Flashcards
Outline the pros and cons of the outer membrane of gram negative bacteria
Note: this whole lecture focuses on gram negative bacteria
PROS
- protection
- restricts access of hydrophobic molecules
CONS
- additional barrier to membrane transport
- no membrane potential across the outer membrane (must rely on diffusion/osmosis for transport)
How do these cells maintain permeability to small molecules in the outer membrane?
Porins - β-barrel proteins spanning the membrane
- form trimeric complexes
- contains constriction loop rich in charged amino acids, acts as filter
Using E.Coli as an example explain how porins can be regulated in response to environmental change
- Cannot be dynamically opened/closed like ion channel
- Regulated by differential gene expression of different sized constriction loops
E.Coli:
Two non-specific porins:
- OmpF: Pore diameter = 1.2nm (high solute flow)
-OmpC: Pore diameter = 1.1nm (reduced solute flow)
- Low osmolarity: OmpF:OmpC = 25:1
- High osmolarity: OmpF:OmpC = 1:15
- prevents excessive solute accumulation which could disrupt osmotic levels in cell
Outline the sensing mechanism behind the response of E.Coli to high osmolarity previously mentioned
1) High osmolarity causes conformational change in EnvZ sensor (maybe through membrane tension)
2) EnvZ is autophosphorylated
3) P transferred from EnvZ to OmpR (a response regulator)
4) OmpR activates OmpC transcription and represses OmpF expression
Give a case study of a molecule-specific porin that is used in gram negative bacteria. Outline the mechanism of this.
- LamB porin specifically uptakes maltose
- 3 selectivity loops discriminate for maltose through van de Waals forces + H bonding
Porin is bidirectional, to avoid maltose leaving cell via LamB maltose quickly taken by transporters:
- Post porin maltose bound to periplasmic binding protein MalE
- MalE delivers maltose to an ABC transporter complex at inner membrane (taken to cytoplasm)
Why do gram negative bacteria secrete proteins and what is required to do so?
- Digestive exoenzymes released to breakdown plant structures for infection
- E.g. Pectate lyase and cellulase secreted by Pectobacterium carotovora
- Injection of pathogenic proteins
- Injection of bacterial DNA e.g. A. tumefacians
- Must have transport across both inner and outer membranes
Describe the translocation systems used to secrete most bacterial enzymes
- Sec or Tat translocation systems at inner membrane
Sec: transports proteins in unfolded state (subsequently folded in plant)
Tat: translocates proteins in folded state
- Most proteins then secreted by type II secretion system made of secretin protein
- IM and OM portions form large multi-subunit complex across both membranes
Describe the secretion systems that are used for injecting bacterial proteins into other cells
Type III Secretion System:
- Most common for injecting pathogens
- Structurally related to flagellum
- Plant pathogen subclass uses Hrp proteins to construct pilus
Type IV Secretion System
- Only system to inject DNA as well as protein
- E.g. Agrobacterium tumefacians
What is Quorum Sensing? Why is it useful?
Outline the mechanism using an example
- QS = communication between microbes
- Often induces phenotype responses that benefit a group but not an individual
- E.g. bioluminescence and establishing biofilms
E.g. V. fischeri:
- acyl homoserine lactone (AHL) (autoinducer) signals synthesised in one cell by Luxl
- AHL diffuse freely across cell membranes
- Perceived by receptor (LuxR) in other cell
- Receptor binds to DNA, activating lux operon expression
- Operon promotes bioluminescence and further AHL production
What are microbial biofilms? Why are they useful?
- Community of microbe cells that stick to each other and to a surface
- Cells supported by matrix of extracellular polysaccharide (EPS), protein + DNA.
- Often used for group protection
- Triggered by AHL-dependent signalling
- E.g. In Pseudomonas aeruginosa EPS synthesised by pel genes.
- pel genes expression depends on QS through pelA-IacZ
Give an example of where a single species contains multiple signalling systems.
Describe why it uses multiple systems?
V. harveyi:
3 autoinducers, all target common transcriptional response of biolum. and biofilm production:
- AHLs, highly species specific, intra-species communication
- Al-2s, widespread, interspecies signalling
- CAI-1
(Function at different levels)
Do other species ever interfere with quorum sensing?
Give an example
- Yes, between bacteria and plant-bacteria
- Often to thwart other individuals for personal gain
- E.g. Alfalfa seed exudates contain non-protein amino acid L-canavanine
- Inhibits AHL production of AHLs by Sinorhizobium meliloti
- S. meliloti beneficial symbiosis for N2 sequestering but this prevents too much spread + biofilm formation
