The Functional Biology Of Membrane Biochemistry Flashcards
Archael membranes
- isoprenoid tails (methyl branches)
- ether bond connects head and tail
- G1 glycerol bond links head group
Bacterial membranes
- fatty acid tails
- ester bond connected head and tail
- G3 glycerol links head group
G1/3 bonds
- convey different stereochemistry
- L/R-handed
Archael/bacterial membrane lipids are
Products of different “universal biochemical pathways” encoded by genes with some overlap; some functional overlap
- flip between
- hard to diagnose membrane biochemistry from genomic data
Archael membrane diversity
- phospholipid mixture variation: in response to stress
- some tail-linked phospholipids
- some within-layer phospholipids (Macrocyclic diethers)
- between layer monolayer (tetraethers, Caldarhcaeol)
- Archaeol
Caldarhcaeol
Strengthens membrane
Archaeol
Ancestral archael lipid
Woese model
- deepest branch marks transitioning membrane chemistry
- further transition @ eukaryagenesis
2D vs 3D tree of life
c40 genes corrected for heterogenous rates of sequence evolution support 2D tree
Independent of the preferred root, the origin of eukaryotes still marks
A transition in cell membrane chemistry
Inside-Out Central Problem
- archael membrane engulfing
- eukaryotic membrane is bacterial
Are shirts is membrane chemistry possible?
Are lipid membrane mixtures possible?
Mixed-membrane E. Coli
- genetic engineering of 2x variants
1) adding archael lipids
2) deleting native E. coli AraM gene - AG in cell fraction (% lipid mixture)
Adding archael lipids
- OX native isoprenoid precursor pathways (MEP/DOXP)
- adding ether lipid bilayer (IPTG inducible gene expression control; optimal at 10μm)
AG
- archaetidylglycerol
- diether lipid
- ^ cardiolipin
- < PG and PE native
- c20% lipid mass
Chimaeric E. Coli
1) delayed growth curve
2) elongate cell form
3) aberrant cell division
4) some resistance to chemical and temperature shock (archael structural stability)
5) bulges and shreds
6) viable!
7) optimal?
Confounding chimaeric E. Coli?
- no IPTG effect control
- can initiate E. Coli elongation
Liposome expts:
- compare mixtures of lipid extracts from a range of eukaryotes
- investigate permeability characteristics of different lipid forms
1) load solute/protons/water
2) load fluorescent marker (reports solute conc.)
3) measure permeability (< fluorescence, as substrate diffuses out of liposome)
Observations of liposome expts
- proton permeability < in all archael lipids (proton gradients determine membrane bioenergetics)
- H2O, urea and glycerol permeability < in mixed archael lipids
Propositions from liposome expts
- ^ archael energetics (Robust to extreme environments)
- embellished isoprenoid archael lipids are much less permeable
- pure archaeol diether cB&E
Confounding liposome expts
- permeability traits measured @ high metabolite conc (100-200mM) ; not physiologically/ecologically realistic
- measured transport out of liposomes
- liposomes = smol , highly curved
- few metabolites measured
Is metabolite permeability a mechanistic difference between membrane types?
Combine microfluidics with unilamellar vesicles
Unilamellar vesicles
- more realistic approximation of cell size/ curvature (10μm)
- constructed from chemically synthesised membrane mimics
- all features of archael + bacterial phospholipids
- pure (no cellular contamination)
- do not react mixtures of lipids found in archaea
Methodology
- Load solute / water (chemostasis)
- Load fluorescent marker (tracks vesicle solute gain)
- Measure permeability (change in fluorescence as substrate permeates into vesicles)
- exposure time = 3 mins
- measure in parallel
Unilamellar vesicle observations and
- archael-membranes are selectively permeable to core requirements of central metabolism
- CONTRADICTION
- 6x aas
- 6x sugars
- urea
- 4x nucleobases
Selective archael permeability
- alanine
- aspartic acid
- tryptophan
- glutamine
- glycine
- leucine
- arabinose
- glyceraldehyde
- glycerol
- dihydroxyacetone
- ribose
- deoxyribose
- many could form autogenously in early Earth
What characteristics determine the permeability function?
- create hybrid fusion synthetic membranes w intermediate chemistries in combination
Hybrid membrane obs?
- chain branching and ether linkage have profound implications on membrane permeability to small metabolites
Hybrid membrane concs
- proton and metabolite permeability show different patterns
Implications of hybrid membranes
- transporter proteome evolution
Comparing TCDB transporter repertoire distribution across prokaryotes
- archaea have consistently < across known classes in transports component database
TCDB
Core metabolite
Heterotrophic origin of cellular life
- cell membrane must have evolved before membrane transporter proteins
- this is only viable with an archael membrane
