1
Q
Membrane functions
A
- Compartmentalization (euks): create separate environments for different activities
- Selectively permeable barrier: prevent unrestricted molecule exchange
- Transport solutes: molecule exchange across membrane
- Energy transduction: convert from one form to another
- Respond to external signals: signals travelling from distance/nearby cells
- Scaffold for biochemical activities
2
Q
What are membrane phospholipids composed of?
A
Polar head group, phosphate, glycerol, 2 fatty acid chains
3
Q
Major polar head groups of phospholipids
A
Used for naming
- Phosphatidyl… choline (PC), serine (PS), ethanolamine (PE), inositol (PI)
4
Q
Amphipathic
A
Have both hydrophilic and hydrophobic parts
e.g. phospholipid, cholesterol (sterol), glycolipid
5
Q
Formation of lipid bilayers
A
- Hydrophobic molecules exclude water, clustering together to minimize energy cost of organizing water molecules
- Amphipathic molecules –> conflicting forces –> solved by formation of bilayer (energetically most favourable, spontaneous)
- Lipid bilayers are: closed, self-sealing
- Sealed compartment formed by phospholipid bilayer –> energetically favourable vs planar phospholipid bilayers (edges exposed to water)
6
Q
Movement of phospholipids within membrane
A
- Phospholipids –> constantly moving within leaflet: lateral diffusion (swapping places within leaflet), twisting, turning, rotation
- Intentional movement: flipping to opposite leaflet during membrane synthesis (rarely flip back)
Allows membrane fluidity
7
Q
Factors that affect membrane fluidity
A
- Temperature
- Change in lipid composition that affect alignment of phospholipid tails
Tightly packed tails –> more viscous, less fluid
Freely moving tails –> higher fluidity
8
Q
Interaction between temperature + lipid composition
A
Membrane fluidity will change if:
- temp changes, lipid composition stays constant
- lipid composition changes, temp stays constant
9
Q
Transition temperature (Tm)
A
Temp at which a membrane transitions b/w fluid phase + gel phase
10
Q
If temp above Tm
A
Membrane ‘melts’ –> lipids move more freely, rotationally, laterally within leaflets
11
Q
If temp below Tm
A
Hydrophobic tails pack together (as if cold, huddling together) –> membrane gels –> incompatible with life
12
Q
Tm + membrane fluidity
A
Cells must maintain fluidity within a relatively narrow range even w/ changes in environmental temp
Usually deal with too cold temps
13
Q
Factors that affect Tm (fatty acids)
A
- Altering length of fatty acid chains
- Longer chains –> more interactions b/w fatty acid tails –> tighter packing –> less fluid at a given temp, higher Tm, higher temp to melt - Altering saturation of fatty acids –> # of db
- More db (more unsat) –> less packing –> more fluid at given temp, lower Tm, lower temp to melt
- Membrane phospholipids typically have 1 sat fatty acid and 1 w/ 1+ db (unsat)
14
Q
Factor that affects Tm (sterols)
A
- Altering amount of sterol (e.g. cholestrol) - (can be up to 50% of membrane lipid in animal cells)
- Cholesterol acts as “buffer”, inhibiting phase transitions when temp changes
- Higher cholesterol at cool temps –> more fluid
- Higher cholesterol at warm temps –> less fluid
15
Q
Examples: variability of membrane fluidity between organisms (within life compatible range)
A
- Organisms from warm climates –> membranes near melt
- Organisms from cold climates –> membranes near gel
16
Q
Mechanisms of membrane fluidity regulation in living cells
A
Homeo viscous adaptation (whole organism level): maintaining fluidity at temps low enough for potential gelling by altering membrane composition
Dealing with low temps:
- Shorter fatty acid chain length (e.g. enzymes cut C18 –> C16)
- Increase # db (= decrease sat): (e.g. desaturase enzymes triggered by low temps)
17
Q
Membranes of archaea vs eubacteria + eukaryotic
A
Both: glycerol, phosphate, 2 HC chains
Archaea: branched ISOPRENE chains (instead of fatty acids) ETHER-linked to L-glycerol
- Allow extremophile archaebacteria to not suffer from membrane breakdown
Eubacteria/Euk: fatty acid chains ESTER-linked to D-glycerol
18
Q
Ways proteins can associate w/ membranes
A
Integral:
- Transmembrane (across entire membrane, both leaflets, external parts outside membrane: alpha-helix, beta sheet
- Monolayer-associated: 1 leaflet
- Lipid-linked: linked to membrane by a lipid
Peripheral:
- Protein-linked: fully outside membrane, linked by integral protein
19
Q
Integral protein association w/ membrane
A
- Hydrophobic R groups allow protein to be located in hydrophobic environment (hold orientation in membrane) since the backbone of protein is always hydrophilic (polar)
- Polypeptide chains usually cross membrane as alpha-helices
20
Q
Formation of hydrophilic channels
A
- From several alpha-helices
- Hydrophobic parts hold protein in membrane
- Hydrophobic parts (very polar) form pore that water and small soluble molecules can flow through
21
Q
Formation of Porins
A
- From beta pleated sheets
- Common in outer membranes of gram-negative bacteria + endosymbiont derived organelles
- Not common in animals
- Very common in bacteria
22
Q
Movement restriction of membrane proteins
A
Restriction of movement by:
- Cytosolic protein
- Extracellular protein
- Identical/non-identical proteins of separate cells interact
- Large membrane protein prevents flow of membrane proteins (e.g. tight junction)
23
Q
Membrane protein distribution in epithelium
A
- Tight junctions create 2 different domains in the membrane, prevent protein movement
- Epithelial cells: line tubular + spherical organs (can bring things in/move things out)
24
Q
Apical domain
A
Domain on top section of cell
25
Basalateral domain
Domain on bottom and side (separated from apical domain)
26
Membrane glycoproteins
- Membrane proteins are coated with sugars on the extracellular side of the membrane
- "Glyco calyx" --> sugar coat(ed)
27
Preservation of membrane symmetry during transport process (sugars)
- Sugar added to protein in Golgi always on non-cytosolic side
- First in the Golgi lumen, then inside the vesicle, then the extracellular side of membrane
28
What is the most extensive membrane compartment?
The ER, especially rough ER
29
Where is new membrane added in a cell?
The ER
30
Role of ER in membrane assembly
Free fatty acids in the cytosol --> catalyzed by enzymes bound to cytosolic side of ER --> new phospholipids added to cytosolic side --> scramblases transfer random phospholipids to other leaflet --> membranes w/ "scrambled" phospholipids emerge from ER
31
Role of Golgi in membrane assembly
Membranes with evenly distributed phospholipids arrive from ER --> flippases selectively move PS and PE to cytosolic leaflet (phospholipids no longer symmetrically distributed) --> membrane asymmetry maintained from this point on
32
Asymmetrical composition of membrane leaflets
- Cytosolic: PS, PE, little bit of PI,
- Non- cytosolic: PC, SM, sugars
- Both: cholesterol equal
33
What does the appearance of PS in outer leaflet indicate?
The cell is going to die
34
Human RBC as model organisms for plasma membrane
- Best understood plasma membrane
- Cells are inexpensive, available in large numbers
- Already present in single cell suspension (don't have to break it down out of tissues)
- Simple: no nucleus, ER, mitochondria, lysosomes (very pure reps of plasma membranes)
- Purified intact plasma membranes --> prepared by producing RBC "ghosts"
35
Membrane transport overview
- Allow passage of certain substances in/out (gases, ions, nutrients, waste)
- Lipid bilayers tend to block passage of polar (water-soluble) molecules
- Can enter by: passing directly, transported by membrane proteins (carriers, channels), engulfed by cell (avoiding passage through membrane)
36
Diffusion
- Dissolved solutes in constant random motion
- Movement high to low concentration
- Solutes spontaneously spread out (increase entropy) until concentrations = in all regions --> no NET flux
37
Semi-permeable membrane
Lets some solutes through and blocks others
38
Osmosis
- Diffusion of water (across semi-permeable membrane) from low solute conc. to high to equalize conc. on both sides
- Outcomes: conc. of water + total solute = on both sides as long as water is allowed to cross --> no NET movement of water
- Solute that CANNOT cross the membrane draws the water across
39
Osmotic pressure
Force opposing the force of water moving from high to low conc. of solute
40
Osmosis + cells
- Water constantly moving through cell membrane in both directions
- If "osmotic tone" (conc. of osmotically active particles) = inside + outside cell --> fluids are isotonic
- If total solute conc. changes on either side (usually outside), net movement of water towards fluid w/ higher solute conc.
41
Cell in hypotonic solution
Net water gain --> cell swells (can lead to hypotonic lysis (pop))
42
Cell in hypertonic solution
Net water loss --> cell shrinks
43
Cell wall in hypotonic solution
- Net water gain
- Turgid (normal) --> rigid cell wall limits increase in size
44
Cell wall in isotonic solution
- No NET water flux, less water coming in than normal
- Flaccid --> wilted, shape changes
45
Cell wall in hypertonic solution
- Net water loss --> plasmolyzed
- Rips cell away from cell wall
46
Turgor pressure
Pressure of cell contents against cell wall in plant + bacterial cells
47
Strategies for maintaining osmotic balance
Osmoconformers: adjust internal salt concentrations to match environment (marine organisms)
Osmoregulators:
- contractile vacuoles --> periodically pump out water (single celled euks --> lack cell walls)
- Terrestrial --> circulate fluid isotonic to cystoplasm throughout body (humans --> blood plasma + extracellular fluid)
Turgor: most plants --> hypotonic to environment --> water pulled into cell --> presses membrane out to cell wall
48
What molecules can pass directly through membranes?
- Depends on size, polarity, charge
- Small nonpolar molecules (conc. gradients): O2, CO2, N2, steroid hormones
- Small uncharged polar molecules (H2O, ethanol, glycerol)
- Larger uncharged polar molecules (AAs, glucose, nucleosides) (negligible ability)
49
What molecules cannot pass directly through membranes?
- Larger uncharged polar molecules (AAs, glucose, nucleosides)
- Ions (H+, Na+, K+, Ca2+, Cl-, Mg2+, HCO3-)
50
Membrane transporter proteins
Carrier protein (shuttle) and channel protein (tunnel) --> both passageways for particular classes of molecules, most are multipass
51
Carrier proteins (shuttle)
- Changes shape to allow transport
- Molecules must fit particular binding site
- 1 molecule at a time
- e.g. Revolving door
52
Channel protein (tunnel)
- Ion channels
- Detects size + charge, as long as channel open --> anything w/ that size/charge can pass
e.g. Open double doors
53
Ion channels
Function: when open, allow movement ('conductance') of Na+, K+, Ca2+, Cl- down their gradients
Critical in many cell activities:
- cell volume regulation
- formation + propagation of nerve impulses
- secretion of substances into extracellular space
- muscle contraction
Features:
- Discriminate charge + size
- Highly selective
- Much faster than carriers (1000x)
- Bidirectional
Ion flux det. by electrical + concentration gradients (electrochemical)
54
Electrochemical gradients driving ion movements
- Voltage + conc. gradients work in same direction --> large flux to lower conc. area
- Voltage + conc. gradients work in opposite directions: net flux changes in different circumstances (e.g. K+)
55
Chemical gradient
Conc. inside vs outside
56
Electrical gradient
Whether it is being attracted across membrane (by oppositely charged molecules) or repelled (by "like" charges)
57
Ion selectivety
- One side determines what moves through (if ion fits), acts as selectively filter
- Other side can be open (e.g. bacterial K+ channel) or closed
58
What determines whether ion channels are open or closed?
- Voltage-gated: charge (reversal) effects protein shape
- Ligand-gated (extra/intracellular ligand): binding of signal changes protein shape
- Mechanically-gated: something like pressure opens the channel
59
How do carrier proteins mediate facilitated diffusion?
- Binding of solute at specific site temp. changes shape of protein
- Solute moves down conc. gradient so carrier protein facilitates passive diffusion
- Many carriers work in both directions
60
Example of carrier protein
- GLUT1 (glucose transporter on mammalian cells)
- Will move glucose but not fructose, D-glucose but not L-glucose
- High specificity
61
Features of membrane carrier proteins
Similar to enzymes:
- Specificity, saturable (revolving door, can only transport amount that fits)
- Can be inhibited/blocked by substances resembling normal cargo (substrate) (like comp. inhibition of an enzyme)
- Passive (facilitates diffusion)
62
Passive vs active transports
- Passive: down conc. gradient (simple diffusion, channel-mediated, carrier-mediated)
- Active: against conc. gradient, requires energy
63
Active transport
- Using energy (directly/indirectly) to move ions against their gradient
- Transport closely coupled to energy release --> hydrolysis of ATP, absorption of light, movement of electrons
64
Sodium-potassium ATPase
- 1st pump discovered (nerve cells of crab)
- ONLY in animal cells
- Moves Na+ out, K+ in
- Coupled to hydrolysis of ATP --> shape change caused by addition of phosphate group
- 3 Na+ out for every 2K+ in --> electrogenic (contributes to electrical diff, inside cell slightly negative)
- AGAINST GRADIENTS of both
65
Action of Na+/K+ ATPase
1. Na+ binds
2. Pump phosphorylates itself, hydrolyzing ATP
3. Phosphorylation triggers conformational change + Na+ is ejected
4. K+ binds
5. Pump dephosphorylates itself
6. Pump returns to original conformation + K+ is ejected
66
Significance of Na+/K+ ATPase
- Both membrane protein + enzyme
- Present in all animal cells: running pump consumes 1/3 energy produced, major contributor to basal metabolic rate, target of many drugs
- Helps maintain a Na+ gradient (high outside, low inside), used to co-transport other molecules (glucose, AAs)
67
Other examples of pumps
Muscle cells: Ca2+-ATPase
Stomach lining: H+/K+-ATPase
68
Function of ion pumps
Allow cells to concentrate certain substances/set up gradients that can be used to drive other processes
69
Carrier proteins + coupled transport
- Symporter: coupled transport in same direction (e.g. Na+ and glucose/AAs)
- Antiporter: coupled transport in opposite directions (H+ exchanged for Na+/K+ or Na+ exchanged for Ca2+)
70
Glucose-Na+ symport in intestinal epithelium
- Using massive Na+ gradient to cytosol (inward) to transport glucose against its gradient
1. Occluded empty
2. Outward open: Na+ binds to active site and waits for glucose to bind
3. Glucose binds --> occluded occupied
4. Flips open randomly, if inward: Na+ and glucose brought in
5. Occluded empty again
71
Glucose transport across gut lining
1. Na-glucose symporter
2. Glut2 facilitated diffusion (uniport, down conc. gradient)
3. Na+/K+-ATPase --> primary active transport (uses ATP)
72
General features of coupled transport
- Membrane carrier protein used driving force of ion (Na+) moving DOWN its gradient to move a solute (small molecule, ion) across the membrane even AGAINST its gradient
- Gradient for ion created by active transport (primary, using ATP)
- Coupled mediated transport (also known as secondary active transport)
73
Pumps in plant cells
- No Na+/K+-ATPase (animal only)
- H+ pumps instead --> protein driven coupled symport
74
Membrane proteins + gene expression
- Each membrane has own characteristic set of channels and carriers (plasma, lysosomal, mitochondrial)
- Transporters are proteins encoded by genes
- Array of transporters present depends on: genes present in organism, whether or not they are expressed (in that cell/patch of membrane at that time)
- Mutation in widely-expressed membrane transporter can have devastating consequences (Cl- --> cystic fibrosis, K+ --> long QT syndrome, Ca2+ --> malignant hyperthermia)
