Topic 4: Membranes and Transport

Question 1

What is the function of the biological membrane?

Answer 1

1. Define boundaries; compartmentalize organelles
2. Compartmentalization allows unique functions for organelles
3. Regulate movement of molecules
4. Cell-cell communication

Question 2

What is the structure of the biological membrane?

Answer 2

composed of amphipathic molecules, contain both hydrophobic and hydrophilic domains

phospholipids are major constituent of the biological membrane

2 fatty acids attached to a glycerol backbone with a phosphate head

fatty acids, hydrocarbon chains 12-20 carbons long, can be saturated (all C-C) or unsaturated (C=C)

Question 3

What is the basic structure of a phospholipid?

Answer 3

polar head group (phosphate head (-) charged)

C=C causes kink in the chain

two fatty acids attached to 3C glycerol backbone

Question 4

What are three important properties of the biological membrane?

Answer 4

stable and self healing

provides sealed and closed compartments

hydrophobic core = major permeability barrier

Question 5

What is the lipid composition of the biological membrane?

Answer 5

phosphoglycerides/phospholipids

sphingolipids

cholesterol

membrane glycolipids

Question 6

What are phosphoglycerides and phospholipids?

Answer 6

derivatives of glycerol-3-phosphate with 2 fatty acyl chains esterified to glycerol backbone and polar head attached to a phosphate group

Question 7

What are the four different head groups that can attach to the phosphate?

Answer 7

PE: small, NH3 containing head group

PS: amino acid head group

PC: largest head group, most abundant

sphingomyelin

Question 8

What are sphingolipids?

Answer 8

polar head group is derived from sphingosine (amino alcohol with large HC tail)

amphipathic but not as cylindrical as phospholipids therefore don’t form bilayers

larger than phospholipids

most common type: sphingomyelin (forms myelin sheath)

sphingolipids form signaling centers called lipid rafts

Question 9

What is cholesterol?

Answer 9

cholesterol derivatives make up the steroids

basic structure is at 4 ring hydrocarbon

has a single OH giving it amphipathicity

abundant in mammalian cells that acts to modulate membrane fluidity

up to 50% of the total membrane lipid in animal cells is cholesterol

usually found in both membranes of the bilayer

orientates itself in the layer using its single hydroxyl group (polar end) to interact with the polar head group of phospholipid (H-bond)

cholesterol is amphipathic but cannot form bilayers

modulates fluidity at high and low temperatures

Question 10

What are membrane glycolipids?

Answer 10

formed by adding carbohydrate (sugar groups) to the lipids

some are glycerol based, but most are derivatives of sphingosine

glycosphingolipids: glycosylated in ER and Golgi, function in cell attachment and communication

prominent components of brain and nerve cell membranes

blood group antigens within the RBC membrane are glycosphingolipids called A antigen and B antigen

membrane glycolipids face the extracellular space (not cytoplasm)

functions: involved in cell recognition in nervous system, different sugars, different functions

Question 11

Why is there a difference between fluidity and flexibility between membranes?

Answer 11

degree of fluidity and flexibility depends on lipid composition, temperature, structure of tails

Question 12

What is membrane fluidity?

Answer 12

membrane fluidity is crucial for proper permeability and cell function

phospholipids are only held together by hydrophobic interactions therefore they are mobile within the bilayers

3D membrane mobility

lateral movement is rapid and frequent

phospholipids are held together by hydrophobic association

flip flop movement is rare without flippases

Question 13

How is FRAP used to study membrane fluidity?

Answer 13

fluorescence recovery after photobleaching

laser can photobleach the fluor if exposed too long

measure degree of recovery, indicates mobility of components

measure time for recovery, indicates degree of fluidity

Question 14

How does lipid composition influence fluidity?

Answer 14

sphingolipid rich membranes are less fluid therefore larger recovery time

PC is less fluid than PS and PE

cholesterol generally restricts phospholipid movement

Question 15

How does the structure of phospholipid tails influence fluidity?

Answer 15

longer HC tails –> less fluid because more hydrophobic

more C=C –> more fluid because less tightly packed

Question 16

How does temperature influence fluidity?

Answer 16

high temperature = more fluid, shorter recovery

low temperature = less fluid, longer recovery

Question 17

What do the FRAP curves look like for biological and artificial membranes?

Answer 17

biological: slow and incomplete

artificial: fast and complete

Question 18

What do the FRAP curves look like for membranes with cholesterol and without cholesterol?

Answer 18

with cholesterol: rapid

without cholesterol: slower but equally complete

Question 19

What do the FRAP curves look like for membranes in high and low temperatures?

Answer 19

high temperature: fast and incomplete

low temperature: slow and incomplete

Question 20

What is membrane lipid asymmetry?

Answer 20

all biological membranes exhibit an asymmetry across the bilayers

all types of phospholipids are present in both leaflets but they are more abundant on one side (leaflet) compared to the other

asymmetrical distribution of phospholipids

outer leaflet: contains sphingomyelin, PC, and glycosylation
inner leaflet: contains PS and PE

asymmetry may affect membrane curvature because exoplasmic face is less fluid

Question 21

How is membrane asymmetry accomplished?

Answer 21

SER is site of phospholipid synthesis (inserted into bilayer at lumenal face)

flippases flip PE and PS to cytoplasmic side

cytoplasmic face always faces the cytoplasm

Question 22

What are other forms of asymmetry?

Answer 22

lipid rafts: microdomains within the bilayers called lipid rafts

rich in cholesterol and sphingolipids

thicker membrane

less fluid

likely signaling centers

Question 23

What are membrane proteins?

Answer 23

proteins are the functional unit of cell, therefore the functions of the membrane like cell-cell recognition, attachment and communication are mediated by membrane-associated proteins

Question 24

What are the three ways proteins can associate with the membrane?

Answer 24

integral membrane proteins

transmembrane proteins

peripheral membrane

Question 25

What are integral membrane proteins?

Answer 25

must be amphipathic in order to span the hydrophobic core of the membrane cytosolic and exoplasmic space must contain hydrophilic surfaces in order to interact with the water in these domains the membrane spanning domain must have hydrophobic amino acids exposed on the surface of the protein to span the hydrophobic core of the membrane alpha helical membrane segments can be inserted: co-translationally at ER and post-translationally at the nucleus and mitochondria

Question 26

What are alpha-helix containing membrane spanning segments?

Answer 26

~22 hydrophobic amino acids are required to span bilayer some pores formed by alpha helical transmembrane domains alternate polar and non polar amino acids

Question 27

What are beta-barrels are another type of membrane spanning domain?

Answer 27

contain alternating polar and non polar amino acid create polar pore e.q. aquaporin beta barrel allows specific passage of H2O

Question 28

What are peripheral membrane proteins?

Answer 28

do not directly contact the hydrophobic core of the membrane but, rather, associate with integral or lipid-anchored proteins or direct interactions to lipid-head groups can be cytoplasmic or exoplasmic but must associate with a membrane component associate by both covalent and non-covalent interactions interact with integral membrane proteins via non-covalent association (quaternary structures)

Question 29

What are lipid anchored membrane proteins?

Answer 29

bond covalently to one or more lipid molecules hydrophobic segment of attached lipid is embedded in one of the leaflets of the membrane anchors protein in the membrane protein chain does not enter the bilayers

Question 30

What are outer leaflet proteins?

Answer 30

proteins that are linked to extracellular surface (GPI anchor, glycosylated inositol) inositol (PI) is a form of phospholipid head that functions in membrane attachment and cell signalling

Question 31

What are inner leaflet proteins?

Answer 31

synthesized in cytoplasm protein is covalently linked to a fatty acid tail

Question 32

What is trypsin?

Answer 32

trypsin digestion allows the identification of the non-transmembrane segments to discover the topology and orientation of membrane proteins trypsin is a non-specific protease

Question 33

How does each transmembrane protein have a specific orientation?

Answer 33

orientation of membrane proteins is established during their synthesis and incorporation into the membrane transmembrane proteins do not flip flop across the membrane another level of asymmetry is the glycosylation of membrane proteins by the Golgi the glyco- part of glycoproteins is only ever found on the exoplasmic face of the membrane

Question 34

How are transmembrane proteins predicted?

Answer 34

the topology of a particular protein can be predicted using hydropathy plots hydrophobic amino acids are counted and given particular values in a computer a strongly hydrophobic domain suggests a transmembrane segment can predict whether it will be a C-terminal, N-terminal, exoplasmic, or multipass transmembrane segments

Question 35

What are N-terminal exoplasmic proteins?

Answer 35

N terminal signal sequence is highly hydrophobic and must also have a hydrophobic stop transfer sequence

Question 36

What are C-terminal exoplasmic proteins?

Answer 36

no N-terminal signal sequence but single strongly hydrophobic peak

Question 37

What are multipass proteins?

Answer 37

multiple hydrophobic domains

Question 38

What are hydropathy plots?

Answer 38

maps large stretches of non-polar amino acids to help predict alpha helical transmembrane segments, look for broad peaks

Question 39

What is the permeability of the bilayer?

Answer 39

permeable to small, uncharged or hydrophobic molecules relatively impermeable to small uncharged polar molecules most impermeable to large uncharged and polar molecules impermeable to charged molecules (ions) because they form favorable interactions with water

Question 40

How do impermeable molecules cross the bilayer?

Answer 40

simple diffusion: movement across hydrophobic core, even impermeable can leak facilitated diffusion: use of a transmembrane channel or carrier, can be gated --> open or closed during different conditions active transport: movement of an impermeable molecule up its gradient, low concentrations to high concentrations, requires input of energy

Question 41

How are transporters involved in facilitated diffusion?

Answer 41

the role of transport proteins is to provide a path through the lipid bilayer, facilitating the "downhill" diffusion of the polar or charged solutes

Question 42

What are carrier proteins?

Answer 42

changes shape with transport bind solutes on one side of the membrane undergo a "conformational" change transfers the solute to the other side of the membrane believed to shield the polar or charged groups of the solute example: GLUT1 carrier, moves glucose down gradient

Question 43

What are channel proteins?

Answer 43

do not change shape form hydrophilic channels through the membrane passage of solutes without a change in the proteins conformation some are non-specific most are highly selective (e.g. ion channels) rapid transport relative to carrier proteins can be beta barrels or composed of alpha helices example: aquaporins, many beta sheet assemble into a barrel, create hydrophilic pore

Question 44

What is an ion channel?

Answer 44

permeable to specific ion like Ca2+ each formed by integral membrane proteins that surround an aqueous pore highly effective net flux depends on electrochemical gradient

Question 45

What are gated channels?

Answer 45

most ion channels exist in either an open or closed conformation such channels are termed gated

Question 46

What are voltage-gated channels?

Answer 46

conformation state depends on charge difference in ionic charge on two sides of membrane example: K+ channel opens with membrane depolarization channels are very specific to solute negative amino acids at opening repels negative ions gating of this channel is accomplished by a voltage center

Question 47

How do you get ion selectivity?

Answer 47

by negative charge at surface selectivity filter pore size

Question 48

How does the pore open?

Answer 48

voltage gated opens when inside of cell becomes more positive (depolarization) normally inside of cell is more negative at depolarization (+50 mV) at resting, positive charged S4 domain is attracted to inside of cell at depolarization, positive charged S4 is attracted to outside of cell shift = open channel

Question 49

What are chemical gated channels?

Answer 49

conformational state depends upon binding of a particular substance example: acetylcholine acts on outer surface of certain cation channels

Question 50

What is active transport?

Answer 50

movement of a molecule up a gradient allows passage of substance through membrane against electrochemical gradient for that substance active transporter acts by undergoing conformational change upon binding substance movement occurs only in one direction three ways that energy can be supplied for active carriers: ATP hydrolysis, coupled transport, light driven example: Na/K pump, establishes resting potential at -70 mV inside, cell constantly pumps 3 Na+ out and 2 K+ against concentration gradient for each ATP consumed

Question 51

What is coupled active transport of existing ion gradients?

Answer 51

movement of two molecules simultaneously, where one molecule moves using the energy gradient from the other molecule symport: two solutes are moved in the same direction antiport: two solutes are moved in opposite directions can couple the resting potential with active transport of another molecule example: use Na+ gradient as a source of energy for active transport of amino acids or sugar