Protein targeting and secretion Flashcards
Where might proteins need to go?
Cytoplasm - nucleoid proteins, ribosomal proteins, metabolic enzymes
Cell membrane - transporters, sensors, motility proteins
Periplasmic space - components of respiratory, sensory and transport systems, some degradative enzymes
OM - porins
Extracellular space - degradative enzymes, toxins
What are signal peptides?
Proteins targeted into or through the cell membrane, often have N-terminal signal sequence, 15-30 aa long
Mix of hydrophobic and +vely charged residues
Usually cleaved off after transport to leave mature N-terminus
What are two common signal peptides?
Sec
Tat - 2 arginines close together
Quite similar
How do integral membrane proteins get into the membrane?
Often have multiple membrane-spanning alpha helices
Signal peptide recognised by SRP (signal recognition particle)
Translation pauses while ribosome directed to SRP receptor FtsY
Protein inserted into SecYEG pore and extruded while synthesised: co-translational
Hydrophobic segments cause protein to escape sideways into membrane
How does Sec transport into the periplasm work?
Signal peptide recognised by chaperones like SecB/Trigger Factor to prevent folding, or directly by SecA
Signal peptide inserted SecYEG: post-translational secretion
Unfolded protein poked through pore using ATP hydrolysis via SecA and maybe also proton gradient via SecDF
When signal peptide enters periplasm it’s cleaved to leave mature N-terminus
How does Tat transport of fully folded proteins work?
Twin arginine transport
Transports fully folded proteins
Signal peptides have 2 arginines, RR, which bind TatC
Proteins fold completely and are directed to TatABC complex
Transient pore forms, numbers of TatA subunits depends on protein size
Protein pushed into the periplasm
How does protein insertion to the OM occur?
Porins, some components of high-affinity uptake systems and motility systems
‘Beta-barrel’ structure with 8-26 antiparallel beta strands
Sec secretion, then bind SurA or Skp chanperone
Insertion into membrane aided by BAM (beta barrel assembly machine)
BamA is the core, BamB, C, and D support
How many protein export/secretion to the medium systems are there?
7
Some start in cytoplasm, some in periplasm following Sec or Tat secretion
How does Type I secretion work?
Related to drug efflux proteins
Each target protein has its own transport system
Transported directly from cytoplasm to medium in single step
Membrane fusion protein interacts with OM protein, TolC, to bring 2 membranes together
Eg. E. coli hemolysin, HlyA transported by ABC transporter HlyB with membrane fusion protein HlyD and TolC
Drug efflux proteins like AcrAB act in similar way, incl interaction with TolC
How does Type II secretion work?
Main terminal branch of General Secretory Pathway
Related to Type IV pili in twitching motility
Exports many different proteins
Targets enter periplasm by Sec or Tat
targets recognised by Type II secreton - involves at least 14 proteins
Protein D forms ring in outer membrane
Protein G ‘piston’ pushes target protein out
How does Type III secretion work?
Related to flagellar macinery
Hollow filament needle complex/injectisome formed extending into medium
Target proteins transported from the cytoplasm through the needle complex into the membrane or cytoplasm of a host cell
Eg. Enteropathogenic E. coli (EPEC) transports Tir into intestinal cell membranes, then binds to it via intimin
Can be used to make silk in bacteria
How does Type IV secretion work?
Related to sex pili used in bacterial conjugation
Capable of transferring DNA and proteins
Eg. Agrobacterium tumefaciens transports T-DNA and Vir proteins To modify plant cell biochemistry
Formation of tumours and secretion of small molecules
Widely used in genetic engineering of plants
How does Type V secretion work?
‘Autotransporters’
Related to OM proteins
Beta barrel domain inserted into membrane as usual but passenger domain pushed through pore
May remain surface-displayed or may cleave itself off with protease domain and released
How can Type V secretion be repurposed?
Can fuse enzyme like alginate to passenger domain - self cleaved
Breaks down alginate into cell
