PM and Cell wall Flashcards
How is bacterial PM composed?
50% lipid (among that 50% is phospholipids), 50% protein.
Bacteria:
2 Unbranched (Linear) FAs with ester linkage to glycerol and 1 phosphate.
PM not rich in sterols,
How is archaeal PM composed?
Branched isoprenoid chains with ETHER linkage to glycerol + 1 phosphate.
Unsaturated.
= Branched, unsaturated, Fatty alcohols!
How can bacterial PM lipid content differ?
Phosphatidic acid can be conjugated with Serine, Choline or Ethanolamine.
To form:
Cardiolipin (CL)
Phosphatidyl-ethanolamine (PE)
Phosphatidylglycerol (PG)
Gram -ve bacteria have more PE
Gram +ve have more PG and CL.
How can lipid content affect the charge of bacterial PMs?
Gram +ve have more PG and CL lipids
Gram -ve have more PE lipids.
PG is negatively charged, whilst PE is neutral.
Therefore, Gram -ve have more neutrally charged PE than Gram +ve which have more negatively charged PG
= Gram +ve have more negative PM.
How does Archaeal and bacterial PM differ>
Ether linkage of glycerol to 2 branched, isoprenoid, unsaturated Fatty Alcohols.
Some parts of archaeal PM is monolayer, where has very long fatty alcohol chains.
= Confers local lower fluidity and reduced permeability.
The branched, unsaturation of isoprenoid provides resistance to dissociation in hyperthermophilic archaea.
How does PM change with respect to temperature?
Unsaturation?
Lipid movements - rotation, chain rotation, lateral diffusion or flip-flop
Each membrane has transition temp - 50% of PM is fluid-like.
Longer chain FAs confer higher transition temperature.
Unsaturation gives lower transition temperature.
Archaeal PM have unsaturation, but can have v. long chain Fatty aclohols.
Are sterols found in bacterial PM?
Archaea?
MOST bacteria lack sterols.
except Spirochetes, Mycoplasma… = MOLLICUTES
Bacteria can have rafts - containing HOPANOIDS.
Archaea have no sterols/hopanoids.
How can sterols/Hoppanoids affect PM fluidity?
Sterols and hopanoids can act as fluidity buffer molecules - serve to rigidify membrane portions.
Especially important when lacking cell walls.
- MOLLICUTES - contain sterols and lack a cell wall.
What are extrinsic and intrinsic proteins?
Intrinsic proteins can be TM or anchored in PM.
= Strongly bound to PM, with TM proteins extending across 2 faces of envelope.
= NEEDS DETERGENT AND ORGANIC SOLVENTS TO REMOVE
= Lipidation, or transmembrane alpha helices/beta barrels…
Extrinsic proteins are weakly bound to PM, and can be only present on of 2 sides
= can have hydrophobic loops, electrostatic binding and in plane alpha helices to anchor..
What is the function of PM?
Highly regulated barrier of permeability and osmosis.
Transport systems for solutes like nutrients and ions.
Involved in energy generation - as bacteria lack mitochondria - can have respiratory/photosynthetic transport systems - ATP synthases in all, but maybe not always ETC.
Coordinate cell wall synthesis.
What antibiotics target the PM?
Bactericidal and detergent-like antibiotics
Polymixins - effective against both -ve and +ve
Daptomycin - against Gram +ve membranes which are enriched with PG
What are the obligatory structures in Bacteria and ARchaea?
Gram +ve = PM, thick peptidoglycan cell wall.
Gram -ve = PM, thin peptidoglycan cell wall, outer membrane
Mollicutes - PM
Archaea:
Gram +ve = PM, thick pseudomurein cell wall
Gram -ve = PM, and outer membrane.
What facultative structures cna be foundd in bacteria and archaea?
Gram +ve = can have an S layer and Polysaccharide capsule.
S-layer = glycoproteinaceous surface monolayer…
Gram -ve = S layer and capsule
Mollicutes = capsule
Archaea do not have capsules, but can both have S layer.
How does Gram staining work?
Crystal violet enters cytoplasm.
Mordant fixes colour
Alcohol is decoloriser, passing into cell to remove crystal violet.
Thick peptidoglycan cell wall blocks alcohol entry in Gram +ve
= purple
Add Safranin counterstain = pink Gram -ve
What can be found in bacterial cell wall?
Gram -ve:
PM, then lipoproteins can be found in periplasmic space.
Thin peptidoglycan layer
Outer membrane, then LPS with porins.
Gram +ve:
PM, with lipoteichoic acids and teichoic acids, with lipoprotein.
Thick peptidoglycan layer.
What is the composition of peptidoglycan?
N-acetylglucosamine (NAG),
N-acetyl muramic acid (NAM)
Alternating NAG-NAM glycans joined with Beta 1,4-glycosidic bonds.
Glycan chain NAMs are linked with lateral, tetrapeptide chains and peptide interbridges or Interpeptide bonds.
How are glycan chains linked in Gram +ve or Gram -ve chains?
Glycan chain NAMs are linked with lateral, tetrapeptide chains and peptide interbridges or Interpeptide bonds.
Gram -ve bacteria like E.Coli have Interpeptide bonds.
DAP and D-ALA of NAM tetrapeptide chains.
Gram +ve bacteria have INTERPEPTIDE BRDGES
Additional glycine residues on NAM link to Lysine of other NAM.
What is the general outline of the stages of Cell wall synthesis?
Cytoplasmic stage:
Synthesis of precursors, UDP-NAM-Pentapeptide and UDP-NAG.
Membrane stage:
Transport across PM with Lipid transporter
Lipid II-NAG-NAM-pentapeptide.
Parietal stage:
Bind to existing peptidoglycan.
Formation of inter-peptide bridges, with loss of terminal Alanine from NAM-pentapeptide.
How are peptidoglycan chains assembled in the final stage of cell wall synthesis?
Parietal stage:
Cross-linking of peptidoglycan chains occurs via PBPs
Penicillin-binding proteins.
PBPs are ALL transpeptidases.
(some can bind NAG-NAMs are transglycolyases)
During transpeptidation, terminal D-Ala is removedto form tetrapeptide, linking D-ala and DAP to form interpeptide bond (Gram -ve)
What classes of antibiotics target cell wall?
Beta lactams:
Penicillins, cephalosporins, carbapenems, monobactams….
Glycopeptides - Vancomycin, teicoplanin
Fosfomycin
= ALL BACTERICIDAL!!!
What is the MOA of beta-lactams?
FIxation to PBPs - inhibition of transpeptidation.
= Prevents cross-link formation during terminal, parietal stage of cell wall synthesis.
How do glycopeptides work?
Against cell wall synthesis.
= Vancomycin, teicoplanin
Inhibition of peptidoglycan synthesis.
Formation of complex with D-ala-D-ala peptidyl residues of peptidoglycan precurssors after emerging from PM.
What is MOA of Fosfomycin?
Prevent formation of NAG:
Inhibition of Pyruvate-NAG transferase.
= Prevents NAG precursor synthesis and therefore peptidoglycan synthesis,
What is lysozyme?
Lysozyme is an acid hydrolase - hydrolyses Beta-1,4 glycosidic bonds between NAG and NAM in peptidoglycan chains.
Present in secretions like tears, mucus, saliva.
Why is the cell wall an important antibiotic target?
Provides mechanical resistance to osmotic pressure.
= In a hypotonic solution, osmosis into cell will lead to increased osmotic pressure.
Maintenance of cell shape/morphology.
= Rod-shape in E.coli can be lost when treated with penicillins.
= Morphology important to preventing amoeba digestion.
-Fixation of bacteriophage and adhesion in Gram +ve.
What are specific components to Gram +Ve cell wall?
THICK peptidoglycan layer.
Teichoic and Lipoteichoic acids:
NO LPS
May have a facultative polysaccharide capsule and S-layer.
What are Teichoic acids?
Gram +ve bacteria.
Polyalcohol of glycerol and ribitol intersperesed with phosphates, which are conjugated by AA residues.
E.G Glucose, NAG, D-ala
The attached residues can vary between bacterial speciesi.
Becomes Lipoteichoic acid when a lipid moiety is attached at extremities.
What is the function of Teichoic acids?
Surface adhesion - to biofilms, to host cells.
Antigenic properties - can be recognised by macrophages.
Anchorage of peptidoglycan.
Regulation of extracellular cation concentration (as has phosphates)
What are specific components to Gram -ve bacteria cell wall?
OUTER MEMBRANE = phospholipid bilayer.
Braun’s Lipoproteins.
LPS
Porins
How is Gram -ve bacterial cell wall composed?
Braun lipoproteins help to anchor the cell wall with the outer and inner membrane.
(as thinner than Gram +ve, less sturdy).
Porins - enable passive transport, antibiotic entry…
LPS on outermembrane.
Thin peptidoglycan layer.
What is the structure of LPS?
Lipid A, with a core polysaccharide and then a Lateral chain called O-antigen.
Lipid A anchors into outer membrane, contains fatty acids, NAG and phosphates.
= 6 FA chains.
(ENDOTOXIN!!!)
Core polysaccharide, contains some sugars and KDO - consistent.
O-antigen - variable long chain of sugars.
= Recognised by immune system.
What is the function of LPS?
Lipid A acts as an endotoxin.
Component of outer membrane.
O antigen is recognised by macrophrages.
Can be recognised and fixated to by Bacteriophages.
Acts as adhesins during biofilm formation.
Lipid A activates TLR4 -
Is there porin diversity?
Porins can diffuse alcohols, sugars, nutrients etc.
Import siderophores - Iron fixation.
Act to efflux antibiotics, toxins etc.
ADHESION
How can transport across both membranes of Gram -ve be coupled?
Maltose serves as example.
Maltose diffuses through LamB porins in outer membrane.
MalE attachment for periplasmic transport.
Active transport, using ATP hydrolysis, via Mal-FGK.
Also glycerol Active Transport too…
How can cell wall weight differ between Gram +ve and Gram -ve?
Gram +ve cell wall acounts for 40% of dry weight.
Around 10% of dry weight.
What bacteria do not have a cell wall?
NOT PRESENT IN MOLLICUTES.
E.G mycoplasma, phytoplasma.
Instead, some may have facultative polysach. capsule.
Would appear pink (gram -ve) in Gram staining.
Evolved to lose Cell wall from Gram +ve bacteria.
= RESISTANT TO penicillins and lysozymes etc.
What is the structure of Archaeal cell walls?
Archea always have an S-layer above cell wall.
Can be Gram +ve or Gram -ve.
BUT NOT PEPTIDOGLYCAN.
What is Gram +ve archaea?
PM, then a layer of Pseudomurein, then S-layer.
Psuedomurein layer anchors S-layer, because Gram +ve S-layer are not anchored to PM/TM.
= Stains purple in Gram +ve
What is Gram -ve archeae?
Large PM anchored glycoproteins form S-layer.
Glycoproteins are TM, with globular heads forming S-layer.
- Forming a monolayer.
S-layer self assembles from monomers to form a almost crystalline structure.
= RELATIVELY RESISTANT + ABLE TO COLONISE HOSTILE ENVIRONMENTS>
What is the S-layer in Archaea?
S-layer self assembles from monomers to form a almost crystalline structure monolayer.
= RELATIVELY RESISTANT + ABLE TO COLONISE HOSTILE
In Gram -ve, glycoproteins are large, anchored in PM. But in Gram +ve, they are anchored to pseudomurein layer.
What is Pseudomurein?
Acetyl-alosaminuronic acid and acetyl-glucosamine
linked with Beta, 1-3 glycosidic bonds.
Forming a glycan chain.
Glycan chains are linked through tetrapeptides and a peptide bridge.
What is role of archaeal envelope?
Mechnical resistance ot osmotic pressure.
Maintenance of cell shape and morphology.
Surface adhesion.
Provide some resistance to hsotile environemtsn as S-layer forms cyrsalline-like monolayer.
