Membranes Flashcards
(157 cards)
1
Q
What are the basic functions of membranes?
A
Form boundaries. Interface between the cell and its environment. Signalling. Controls entry and exit. Site of specialised chemical reactions. Permit vectorial reactions.
2
Q
Give an example of control of entry and exit of materials in cells.
A
In plant cells, external potassium concentration is 0.1 - 1 mM, while cytosolic potassium concentration is 80mM.
3
Q
Why are endomembranes key for organelle specialisation?
A
Allow different environments to be maintained inside the organelle.
4
Q
Where in membranes are polysaccharides found, and what percentage of the membrane do they make up by weight?
A
Found as part of glycoproteins and glycolipids on the external surface of the membrane. They make up about 10% of the membrane by weight.
5
Q
What is the most common type of phospholipid in membranes?
A
Phosphoglycerides.
6
Q
Describe the structure of phosphoglycerides.
A
Formed around the glycerol. C1 and C2 are bonded to two (usually) different fatty acids by ester linkages. C3 forms and ester bond with a phosphate group. The phosphate group can then form an ester linkage with a charged head group (e.g. an amino acid or an alcohol).
7
Q
What is the nomenclature for phosphoglycerides? Give an example.
A
Phosphotidyl + charged group.
Phosphotidyl choline.
8
Q
Give a type of lipid beginning with s.
A
Sphingolipid.
9
Q
Describe sphingolipids.
A
Formed around the long-chain, nitrogen-containing alcohol sphingosine. The amino group on sphingosine forms an amide linkage with a long-chain fatty acid to form a ceramide. The OH group on C1 of the ceramide forms an ester linkage with a phosphate group, which then binds to a polar head group. Alternatively, the OH group can form as ester bond with a polysaccharide.
10
Q
Give one example of a sphingolipid that is a phospholipid and one example that is a glycolipid.
A
Ceramide 1-phosphoryl choline (sphingomyelin).
Monogalactosyl ceramide.
11
Q
What are glycolipids?
A
Lipids containing a carbohydrate that can range from a monosaccharide to a branched oligosaccharide.
12
Q
What are steroids?
A
Lipids containing 4 interconnecting rings of carbon atoms with varying numbers of double bonds and different side groups.
13
Q
What are sterols?
A
A class of steroids containing an OH group at one end and a non -polar hydrophobic chain at the other.
14
Q
Why do non-polar fatty acid tails self-associate?
A
This reduces the surface area of the hydrophobic regions that are in contact with water, reducing the ordering of water molecules around the fatty acid tails, thus increasing entropy, making this arrangement thermodynamically stable.
15
Q
What forces, other than hydrophobic interactions, stabilise the packing together of fatty acid tails?
A
Van der Waals forces.
16
Q
What forces stabilise the packing together of the polar headgroups?
A
Ionic bonds and hydrogen bonds.
17
Q
Why is there no repulsion opposing close packing of lipids?
A
At physiological pH, most lipids are zwitterions, so there is no charge repulsion opposing close packing.
18
Q
What is formed when the edges of the bilayer are spontaneously brought together? Why does this happen spontaneously?
A
A liposome.
Reduces the surface area of hydrophobic fatty acid tails exposed to the aqueous medium.
19
Q
What determines the degree of curvature possible in a membrane?
A
The glycolipid content - the greater the proportion of glycolipids in the membrane, the higher the degree of curvature possible (e.g. thylakoid membranes).
20
Q
At low temperatures, what state is the membrane in, and what does this mean?
A
It is in the ‘gel state’. This means that the hydrocarbon tails are packed tightly together and have restricted motion.
21
Q
What is the phase transition temperature for a membrane, and what happens to a membrane when it is reached?
A
It is a peak of heat absorption at which the hydrophobic interior becomes more fluid. The membrane is then said to be in the ‘liquid crystal state’
22
Q
At high temperatures, what happens to the membrane?
A
The forces holding it together are interrupted and the bilayer is dispersed.
23
Q
The more varied the composition of the bilayer, what is the effect on the phase transition temperature?
A
It has a broader range.
24
Q
Shorter fatty acids have a lower melting point. What is the effect of this on membranes with a high proportion of short-chain fatty acids?
A
They undergo phase transition to a liquid crystal state at a lower temperature.
25
Why do membranes containing a lot of unsaturated fatty acids have a lower phase transition temperature?
Unsaturated fatty acids can't pack together very well because the double bonds cause the chains to kink. This means they have a lower melting point as the Van der Waals forces and hydrophobic interactions between them are less effective.
26
In eukaryotes, what lipids are used to regulate membrane fluidity?
Sterols.
27
Explain how sterols control membrane fluidity.
They intercalate between phospholipids with the polar hydroxyl group towards the exterior of the membrane and the hydrophobic steroid ring and tail are to the inside. They restrict the movement of the hydrocarbon chain near the head-group, but disperse the tails. Below transition temperature, this prevents the chains packing together too closely. Above transition temperature it restricts the movement of the hydrocarbon chains near the head-group, preventing dispersal of the bilayer.
28
How do plants, bacteria and poikilothermic animals regulate membrane fluidity?
Through 'de novo' synthesis and modification of fatty acids. For example, at low temperatures, synthesis of unsaturated fatty acids may be increased.
29
What happens to mobility of lipids above the phase transition temperature?
It increases.
30
What are the ways that lipids can move in the membrane?
Lateral movement
Spin about their longitudinal axis.
Flex their hydrocarbon chains.
Flip-flop motion from one half of the bilayer to the other.
31
Why are enzymes required for flip-flop motion, and what are these enzymes called?
They are required because movement from one half of bilayer to the other is thermodynamically unstable and would require a high input of energy. The enzymes that catalyse the reaction are called 'flippases'.
32
Explain some evidence for flip-flop in phospholipids in liposomes.
Phosphoplipids spin-labelled.
At first, all of the phospholipids are spin-labelled so the percentage of the label that absorbs is 100%.
Then add ascorbate, which reduces the labels on the outer layer of the liposome membrane, lowering the percentage of original label that absorbs.
Then remove ascorbate. If some flip flop occurs, then some of the unreduced labels end up on the outside, and some unlabelled phospholipids on the inside.
Add ascorbate again.
If absorbance is reduced again, then flip-flop has occurred, as some of the internal labelled ones slipped onto the outside and were reduced.
33
What is a spin-label?
A spin-label is an unpaired electron that will absorb energy at a specific wavelength in a magnetic field and produce a characteristic absorption spectrum.
34
Can cells make membranes from scratch?
No - they must synthesise membranes by expanding old ones.
35
Where are the enzymes for phospholipid synthesis situated? Which way does their active site face?
They are membrane-bound, and their active sites face towards the cytosol, from which they receive substrates.
36
In bacteria, where are membranes synthesised?
At the plasma membrane.
37
In plants, animals and fungi, where are membranes synthesised?
Cytoplasmic side of the SER or in the Golgi lumen.
38
What is the implication of the position of the active sites of enzymes that synthesise phospholipids?
Lipids are only incorporated into one half of the lipid bilayer, so some must flip-flop to the other half.
39
What is meant by saying that lipid asymmetry is 'NOT absolute'?
Any type of lipid is present in both halves of the membrane, though their amounts may vary.
40
What is meant by saying that protein asymmetry 'IS absolute'?
All copies of any one protein face the same way.
41
What is meant by saying that carbohydrate asymmetry 'IS absolute'?
Carbohydrate moieties on glycolipids and glycoproteins are all situated on the non-cytosolic side of the bilayer.
42
What is the term used when a protein traverses the bilayer multiple times?
Multi-span.
43
How can the parts of a transmembrane (type I) protein that are not embedded within the membrane anchored to the membrane?
May be anchored to it via a covalently bound fatty acid chain that inserts into the lipid bilayer.
44
In what ways can a hydrophobic type II protein be attached to the lipid bilayer?
Covalent attachment directly to fatty acid in the cytosolic half of the membrane, or covalent attachment via an oligosaccharide to a phospholipid (usually phosphatidylinositol) in the outer half of the bilayer.
45
What is the name for proteins that are either transmembrane or anchored to the membrane covalently, and cannot be detached without disrupting the membrane?
Integral proteins.
46
As well as integral proteins, what other type is there, and how do they attach to the bilayer?
Peripheral membrane proteins. They attach through non-covalent interactions with integral proteins in the membrane.
47
Do peripheral membranes require the membrane being disrupted in order to be removed?
No.
48
How can you find out if a protein spans the plasma membrane?
Label vesicles with lactoperoxidase and hydrogen peroxide. Lactoperoxidase catalyses the peroxide-dependent iodination of the protein's tyrosine residues.
At first, since it can't go through the membrane, only external residues will be labelled.
Disrupt the vesicle and repeat, in which case any tyrosine residues inside will also be labelled.
Extract the proteins from both the disrupted and complete vesicles and treat them with proteases.
Perform SDS polyacrylamide gel electrophoresis.
If the protein spanned the bilayer, then the proteins from the disrupted sample would have additional radioactive bands.
49
How must proteins be treated in order to perform electrophoresis?
Must be denatured as their shapes may vary and affect movement by using a reducing agent like mercaptoethanol to break disulfide bonds.
Must be treated with the anionic detergent sodium dodecyl sulfate (SDS) to eliminate charge density as a variable.
50
Why is it important that shape and charge density are removed as variables?
So only the length of the polypeptide chain differs.
51
Describe the cell fusion method that provides evidence for lateral movement in the lipid bilayer.
Make antibodies complementary to the mouse cell membrane proteins and conjugate them to fluorescent dye.
Do the same for human cells, but with a different colour fluorescent dye.
Fuse the cells.
Excite the dye.
At first the different dyes will be on different halves of the cell.
Excite the dye again some time later. The colours will have mixed.
52
Describe the Fluorescence Recovery After Photobleaching (FRAP) method that provides evidence for lateral movement in the lipid bilayer.
Treat cells with concanavalin A, a plant lectin that binds to the carbohydrate moieties of surface glycoproteins, conjugated to a fluorescent dye.
Use a laser to photobleach a small area of the cell.
Time the return of fluorescence to indicate the rate of lateral movement of glycoproteins.
53
If in a test with FRAP, only 55% of the fluorescence is recovered, what percentage of the glycoproteins were mobile?
55%.
54
How can lateral mobility of proteins be reduced?
Attachment to the cytoskeleton.
55
Explain how the 'Band 3' channel protein in erythrocytes, which allows exchange of the anions Cl- and HCO3-, is held in place.
Spectrin dimers join 'head-to-head' to form tetramers.
The tetramers link at the tail end to short actin filaments and to 'band 4.1' proteins. Spectrin is anchored to the cytoplasmic surface of the membrane by molecules of ankyrin, which are bound tightly to the 'band 3' protein.
56
What sort of proteins are spectrin, actin and 'band 4.1'?
Peripheral proteins.
57
What happens when erythrocytes are treated with low ionic strength buffers?
The peripheral proteins, spectrin, actin and 'band 4.1' dissociate from the membrane, causing the cells to lose their biconcave shape, and membrane proteins show lateral mobility.
58
Describe the signal sequence and its position in proteins that are NOT destined to stay in the cytosol.
The signal sequence is 15-30 amino acid residues long and it at the N-terminus of the protein.
59
Once 70-80 amino acids have been polymerised, the signal sequence emerges into the cytosol. What happens from then up until the ER recognises whether it is going to be an integral protein?
The signal sequence is bound by the 54kDa protein component of the signal recognition particle (SRP).
Binding to the SRP slows or even stops protein synthesis.
The SRP binds to an integral SRP receptor in the ER membrane.
There is also a ribsome receptor where the ribosome binds.
Once the ribosome is bound the SRP is released and can be reused.
Loss of the SRP allows protein synthesis to be resumed.
The signal sequence associates with the ribosome receptor, which acts as a tunnel to allow the polypeptide to traverse the membrane as it is synthesised.
As this happens, the ER recognises whether this is going to be an integral protein or not.
60
What happens if the ER recognises a protein to be a non-integral protein?
The protein emerges into the ER lumen and the pore closes behind it.
It is folded by ATP-dependent 'foldases'.
61
Where do ATP-dependent 'foldases' bind?
They bind at exposed hydrophobic surfaces that are usually buried within the folded protein.
62
How are integral membrane proteins recognised by the ER, and what happens after this?
Integral membrane proteins have a hydrophobic 'stop-transfer' sequence. This halts their transfer through the translocation tunnel of the ribosome receptor, leading to the 'fixing' and orientation of these proteins in the bilayer.
63
What is the final modification to a newly synthesised protein?
The signal sequence is cleaved off.
64
What do proteins that are destined to remain in the ER have?
A retention sequence.
65
Which proteins have sorting sequences, and what is the function of these sequences?
Proteins that are destined for export from the ER have these sequences. Their function is to target proteins to specific cellular compartments.
66
Where are proteins with no sorting sequence exported to?
They either end up in the plasma membrane or are excreted.
67
Describe the plasma membrane/ secretory pathway.
Secretory and membrane protein leave the plasma membrane in transport vesicles which bud off from specialised, ribosome-free areas of the ER.
They are delivered to the cis face of the Golgi, then pass through the cisternae of the Golgi in repeated cycles of budding and fusion.
Some of the proteins are 'resident proteins' intended to stay in the ER, and they have a KDEL retention sequence at their C-terminus, so are redirected back to the ER.
Proteins glycosylated in two stages:
1. Sugars making up the basal structure of the carbohydrate moiety are added as one oligosaccharide to one of the three amino acids that can form glycosdic bonds (asparagine, serine and threonine).
2. Addition of more carbohydrate residues to make side-branches occurs with every passage through the Golgi apparatus.
68
What other process can take place to integral proteins in the ER?
The covalent attachment of fatty acids or phospholipids used to anchor the proteins to the plasma membrane.
69
How are components retrieved from the plasma membrane?
The pinching off of small sections of membrane to form vesicles in the cell (endocytosis).
70
How are components integrated into the plasma membrane?
The fusion of a vesicle containing the components in its membrane with the plasma membrane (exocytosis).
71
As well as transporting proteins to and from the plasma membrane, what can vesicles be used for?
Transporting solutes.
72
Describe how endocytosis can transport solutes into the cell.
As the plasma membrane invaginates, it will capture some of the external medium which will then be transported into the cell.
73
What is the endocytosis of solids called?
Phagocytosis.
74
What is the endocytosis of liquids called?
Pinocytosis.
75
Describe receptor-mediated endocytosis.
The solute binds to the extra-cellular domain of a specific receptor protein.
In the first 22 residues of the cytoplasmic C-terminus of the receptor, there is a tetrapeptide sequence that allows the receptor to bind to an extrinsic protein adaptor.
This adaptor then interacts with another cytoplasmic protein called clathrin, which has three heavy and three light chains, forming a triskelion.
The triskelions assemble below the plasma membrane and spontaneously polymerise, forming coated pits of invaginated membrane.
The protein adaptor associates with the free end of the heavy chain of clathrin.
If not already located above the pit, the binding of the solute induces a conformational change that allows the receptor to be captured when it diffuses into the pit system.
Invagination of the membrane continues until it excises to form a clathrin coated vesicle.
In the cytosol the vesicle uncoats to form an early endosome, containing the receptors facing into the lumen, bound to the solute.
The clathrin and adaptors are recycled.
76
How is cholesterol transported in animal cells?
Cholesterol is transported as a low-density lipoprotein (LDL) because it is insoluble. This consists of a core of cholesterol molecules ester-linked to fatty acids, surrounded by a monolayer of phospholipids and unesterified cholesterol.
77
How do LDLs bind to receptors?
There is a specific protein embedded in the monolayer that allows recognition of the LDL.
78
What happens once the early endosome has been formed?
The early endosome fuses with a specialised compartment known as the late endosome or CURL (Compartment of Uncoupling of Receptor and Ligand). The CURL lumen is at a lower pH of 5.5 which causes the solute to dissociate from the receptor. The receptors become embedded in the tubular extensions of the CURL, which bud off to form transport vesicles to take the receptors back to the plasma membrane.
79
What happens to LDL once it has dissociated from its receptors in the late endosome?
It is transported to a lysosome by other vesicles where the fatty acids are cleaved from cholesterol.
80
Explain how exocytosis works.
Fusion of vesicles to the plasma membrane can release solutes to the extracellular matrix and insert components into the plasma membrane.
Different vesicles are targeted to different locations in the plasma membrane by specific protein-protein interactions.
81
Give an example of high exocytotic activity.
In seed germination, specialised aleurone cells exo-cytose alpha-amylase to hydrolyse the seed's starch supply in the endosperm.
82
What is the key difference between transmembrane transport and vesicle-mediated transport?
In vesicle mediated transport, the solute remains at the same face for the duration of transport, whereas in transmembrane transport the solute actually moves through the membrane.
83
What are the features of diffusion through the lipid bilayer?
Passive.
Only small, non-polar molecules can diffuse through at an appreciable rate, although some small polar molecules like water can permeate.
84
Describe the process and principles of passive transport.
The solutes have kinetic energy so move around. They diffuse down chemical potential or electric potential gradients because there is a free energy difference. As diffusion takes place, the difference decreases and entropy increases.
85
In electrochemical gradients, are the electrical and chemical gradients always additive?
No, they can also oppose one another.
86
What does the presence of a protein add to passive transport?
It adds specificity and can affect kinetics.
87
What is the term used to describe passive transport through a protein?
Facilitated diffusion.
88
What is the effect of active transport on the free energy and entropy of a system?
Free energy increases and entropy decreases.
89
What is the equation for Gibbs free energy change for an uncharged solute?
ΔG = ΔG° + RTln([X]i/[X]o)
90
Why is the equilibrium constant for diffusion always 1?
Keq = [X]i/[X]o
91
Why is the standard free energy change (ΔG°) equal to 0 at equilibrium?
ΔG° = -RTln(Keq) and Keq is always 1 at equilibrium. ln(1) = 0, so the expression is equal to 0.
92
What is the equation for the standard free energy change?
ΔG° = -RTln([X]i/[X]o)
93
Given that standard free energy change is zero at equilibrium, what does the equation for Gibb's free energy change simplify to at equilibrium, with an uncharged solvent?
ΔG = RTln([X]i/[X]o)
94
In inward transport, does the internal or external concentration of solute act as the numerator for the expression for the equilibrium constant?
The internal concentration [X]i.
95
When [X]i is less than [X]o, what will be the effect on the Gibbs free energy change, and hence the reaction?
Gibbs free energy change will be negative, so the reaction will be exergonic and occur spontaneously.
96
What is the equation for Gibb's Free Energy change with a charged solute?
ΔG = RTln([X]i/[X]o) + zFEm
Where z is the charge on X,
F is the Faraday constant (96.5 kJ mol^-1 V^-1)
Em is the membrane potential in volts.
97
What assumption do you make to rearrange the equation to find out the membrane potential?
Assume the system is at equilibrium so ΔG is zero.
98
Give the Nernst equation and the conditions at which is applicable.
Em = 58/(zlog([X]i/[X]o))
| When T=293 K.
99
If we know the membrane potential and find that the solute concentrations do not match those predicted by the Nernst equation, what can we assume?
The distribution isn't achieved by passive transport.
100
What coefficient can be used to quantify the lipid-solubility of a solute? Give the equation for it.
Partition coefficient.
| Partition coefficient = amount dissolving in test lipid/amount dissolving in water.
101
What is the general trend with increasing lipid solubility?
Greater permeability.
102
What are two exceptions to the general trend, in that they have a low partition coefficient, but high permeability.
Water and urea.
103
What is an important factor in the permeability of polar or charged molecules?
Size.
104
For small, non-polar and uncharged molecules, what is the factor that determines rate of diffusion across the bilayer? What law is this?
Permeability coefficient and difference in concentration. Fick's first law.
105
What is the permeability coefficient?
Amount permeating the membrane per unit surface area per unit time.
106
How can transport proteins be regarded?
As enzymes.
107
Give some key features of protein-mediated transport.
Can be highly specific.
Can be inhibited.
Rate of transport/reaction far higher than through the lipid bilayer.
Shows saturation kinetics.
108
Is passive transport intrinsically vectorial?
No.
109
Describe the structure and function carrier proteins involved in passive transport.
Usually have about 12 alpha-helices spanning the membrane.
They are very useful for transporting amino acids and sugars.
The solute binds to carriers on the side of the membrane with greatest concentration, inducing a conformational change that prevents any more solute binding and releases the solute to the other side of the membrane.
Once the solute is released, the protein returns to its original conformation and the process repeats.
If the (electro)chemical is reversed, so is the direction of transport.
110
How do Fungi overcome the non-vectorial nature of passive transport to obtain enough glucose? Where else does this process occur?
They phosphorylate glucose as soon as it reaches the cytoplasm, so it is no longer complementary to its carrier protein.
Also occurs in erythrocytes.
111
Describe the structure and function of channel proteins.
Channel proteins transport ions.
Specificity is thought to be given by the surface charge, narrow filter and specific residues lining the aqueous pore.
Exit of ions from the pore is controlled by a conformational gate which gives the channel discrete open and closed conformations.
112
Why are channel proteins thought to be the fastest enzymes known (10^6 - 10^8 molecules per second)?
The smaller conformational changes that occur.
113
Give three examples of how the conformational gate of a channel protein can be controlled.
It may have positively charged amino acid residues that undergo conformational changes in response to changes in membrane voltage.
Phosphorylation by kinases.
The binding of extracellular ligands like cAMP.
114
How can ion transport through channels be measured?
Patch clamp electrophysiology. A blunt glass microelectrode is pushed against the membrane, electrically isolating the patch of membrane with a single channel protein in it. The tiny current that flows when ions flow through the channel can then be measured.
115
Give two examples of when rapid ion transport through channels is needed.
Muscle contraction and nerve transmission.
116
What are ionophores? Give an example.
Bacterial peptides that shuttle within a membrane, and can carry ions through it. They can therefore collapse ion gradients. An example is valinomycin, which can potassium ions in its centre and shuttle them through the membrane as it has a hydrophobic surface.
117
What are the three main ways that water is transported?
Diffusion.
Bulk/mass flow.
Osmosis.
118
Define diffusion in terms of water.
The net movement of water from an area of high energy to an area of low energy.
119
What is the bulk/mass flow of water?
Movement of water in response to a pressure gradient.
120
What is osmosis?
The movement of water through a differentially permeable membrane from a region of high water potential to a region of low water potential.
121
What is water potential?
The chemical potential (J mol^-1) divided by the partial molal volume of water.
122
What is the partial molal volume of water?
The volume of one mole of water (18x10^-6 m^3 mol^-1)
123
What is ψ_s?
Solute/osmotic potential. It describes the effect of dissolved solutes on water potential.
124
What is ψ_p?
Pressure potential. Describes the effects of positive or negative pressure.
125
What is ψ_g?
Describes the effect of gravity and only has an appreciable impact above 5m.
126
Water potential is a relative term. What is it relative to?
It is relative to the water potential of pure water at sea level at standard temperature and pressure (0MPa).
127
What is the general effect of dissolving solutes in water on the free energy of the water?
It decreases, as the water is diluted, which increases entropy.
128
The van't Hoff equation is used to find the solute potential for a non-dissociating solute. Give the equation.
ψ_s = -RTc_s
| Where c_s = solute concentration as moles of solute per litre of water (osmolality).
129
If a solute dissociates, what is changed about the van't Hoff equation?
c_s is multiplied by the number of particles into which it dissociates.
130
What is the effect of negative pressure on water potential?
It decreases it as there is less energy.
131
Why is the effect of pressure on water potential important in walled cells?
It helps to prevent damage to the cell - when water diffuses into the cell by osmosis, the protoplast swells and pushes against the cell wall. This increases pressure, increasing the water potential until it is equal to the water potential of the surrounding solution, which helps to prevent any further osmotic gain of water.
132
What is the state of a plant cell when the protoplast has peeled away from the cell wall?
Plasmolysed.
133
What is a hypotonic solution?
Solution with a higher water potential than the cell.
134
What is a hypertonic solution?
Solution with a lower water potential than the cell.
135
What is an isotonic solution?
Solution with the same water potential as the cell.
136
What are aquaporins?
Channel proteins that allow the passage of water.
137
What is required in active transport to move solutes against their (electro)chemical gradient?
A discrete input of energy.
138
Where does the energy input come from in primary active transport?
Hydrolysis of an energy rich compound like ATP.
139
Where does the energy input come from in secondary active transport?
Energy input comes from the movement of an ion down its (electro)chemical gradient.
140
What are three key features of active transport?
Can occur against a concentration gradient.
It is highly specific.
Shows saturation kinetics.
141
What are the three main energy donors in active transport?
```
Adenosine triphosphate (ATP)
Guanosine triphosphate (GTP)
Inorganic phosphate (PPi)
```
142
What is common to all of the main energy donors for active transport?
They have a high energy phosphate bond, which is hydrolysed by the protein.
143
Why are the phosphoanhydride bonds in ATP so high in energy?
At physiological pH, the hydroxyl groups on the phosphates have lost their proton, leaving negatively charged oxygen atoms. This repels the electrons in the P=O double bond, leading to P having a partial positive charge. The phosphoanhydride bonds between the phosphates msut therefore have enough energy to overcome repulsion between the positively charged phosphate groups, and hence must be high energy.
144
What is the standard free energy change when ATP is hydrolysed?
-31 kJ mol^-1.
145
What is the collective name of primary active transporters that hydrolyse ATP in order to catalyse a transport reaction?
Solute-transporting ATPases.
146
What is the conserved feature of a P-type ATPase?
The formation of a phosphorylated intermediate during the reaction.
147
Give an example of a P-type ATPase and its function.
The sodium-potassium pump.
The binding of 3 sodium to the protein and the phosphorylation on the same internal side of the membrane leads to a series of discrete conformational changes that result in the transference of the bound Na+ to the extracellular surface. The pump is returned to its original position in a series of conformational changes brought about by the binding of two K+ ions on the extracellular side of the membrane. In this way, the hydrolysis of ATP brings about the transference of 3Na+ out of the cell, and 2K+ into the cell.
148
In plant and fungal cells, the sodium-potassium pump is not pre-dominant. What protein is, and what is its function?
Another P-type ATPase that pumps 1 proton per ATP hydrolysed, maintaining a large inside-negative membrane potential and pH gradient.
149
What are V-type ATPases?
Primary active transporters that pump H+ across a membrane. They are multi-subunit enzymes containing an integral (V_0) domain and a peripheral (V_1) domain.
150
How does secondary active transport work?
The free energy released by the movement of a solute down its (electro)chemical gradient can be coupled to energetically unfavourable processes.
151
If the solute being translocated against its own concentration gradient moves in the same direction as the solute that is being passively transported, what is the protein called?
A symporter.
152
If the solute being translocated against its own concentration gradient moves in the opposite direction to the solute that is being passively transported, what is the protein called?
An antiporter.
153
Give an example of secondary active transport.
Glucose absorption in the gut.
The sodium-potassium pump in the basolateral membrane of the cell sets up an Na+ gradient between the outside and inside of the cell, so sodium diffuses down its concentration gradient into the cell through a Na+-driven glucose symport in the apical membrane. The free energy released by this process is coupled to the transport of glucose through the protein against its concentration gradient.
154
What is an F-type ATPase?
A transporter protein that uses the free energy released by passive proton transport to synthesise ATP.
155
What are the two domains of the F-type ATPase?
The integral F_0 region and the peripheral F_1 region.
156
Describe how the F-type ATPase works, with reference to the diagram in your notes.
Protons enter the protein at the c-a interface, disrupting the interactions between the c and a subunits, causing c to rotate. This leads to the rotation of the γε shaft, which rotates 120 degrees anti-clockwise. The F_1 region has a stationary section comprised of three non-catalytic alpha subunits and 3 catalytic beta subunits. The rotation sequentially changes the conformation of the beta subunits from loose to tight to open. In the loose conformation, the subunit binds ADP and Pi. In the tight conformation, there is a high affinity for ATP so it is formed from ADP and Pi. In the open conformation, ATP is released.
157
How do signalling ligands work?
They bind to a specific transmembrane protein receptor, triggering a conformational change that triggers subsequent conformational changes in functionally linked cytoplasmic proteins, triggering the next specific signalling reactions.
