Membrane proteins Flashcards
How are tightly bound membrane proteins solublised?
Use of detergent or organic solvent. Sometimes solutions with high ionic strength
Why are integral membrane proteins difficult to separate from the plasma membrane?
They interact with internal fatty acid tails.
Can be released by agents which compete for these non-polar interactions
How are peripheral membrane proteins bound to lipid heads?
Bound by electrostatic and hydrogen bond interactions.
Can be bound to surfaces of integral proteins
What types of hydrophobic lipid anchor are there?
Acylation
Prenylation
GPI lipid anchor
What is acylation?
Hydrophobic lipid anchor on the inner leaflet of plasma membranes. Same length as hydrocarbons in phospholipids. N-terminal methionine has to be removed to expose glycine. Amide binds to this.
What is prenylation?
Hydrophobic lipid anchor with a thioester link to cysteine on the C-terminus. This pattern is -Cys-a-a-X (a being an aliphatic amino acid). When something attaches to the C-terminus, aax is cleaved.
What are GPI anchored proteins comprised of?
Phosphate, inositol, mannose and ethanolamine
Hydrophobic pore, hydrophilic on the outside
Why aren’t GPI-anchored proteins used for transport across the membrane?
They don’t span the membrane
In membrane channels, why do residues on the outside of the pore tend to have ring structures?
Act as dampers between hydrophilic and hydrophobic residues
Why is hydrogen bonding important in β-barrel proteins?
It is energetically unfavourable for unpaired carbonyls to be in a hydrophobic environment, hydrogen bonding counteracts this
What happens when there is a hole in the plasma membrane?
Everything would diffuse out the membrane and polarity would be lost
Proton gradient would disappear
How is bacteriorhodopsin structured?
7 α-helicies form a tight bundle. These are mainly made up of non-polar residues. Retinal is covalently bonded and absorbs light.
Where is free energy of amino acids in bacteriorhodopsin scored?
α-helix in the membrane interior to water
How can alpha helix structures be predicted?
Balancing free energy transfers of amino acids with hydropathy plots.
Why cannot β-barrel structures be predicted with hydropathy plots?
They have alternating hydrophobic and hydrophilic residues, charges would balance out.
How can membrane permeability be calculated?
(diffusion coefficient X water-membrane partition coefficient) / membrane thickness
What does a basic channel comprise of?
Hydrophobic interface which protects the core from the membrane
Vestibule- where the compound is collected
Narrow pore with a selectivity filter
Describe the structure of aquaporins
Tetramer made of 4 individual subunits, each with its own pore- 6 hydrophobic helices each
Helices are tilted at 30° and twisted into a right handed bundle.
2-fold symmetry axis (inverted topology repeats), indicating ancient gene duplication events
How do aquaporins make sure one water moves through the channel?
Arginine electrostatically repulses protons.
If water moving through is uninterrupted, a proton conducting wire is formed.
NPA motifs in the middle prevent ions moving through
Asn76 and Asn192 form hydrogen bond donors which flip the water dipole oxygen up, hydrogen down. This creates a temporary break, preventing protons moving with it.
What was the first evidence for glucose transporters?
Rate of glucose transport capped after a certain concentration. Transport was shown to be saturable.
Outline the mechanism of GLUT1
- ATP is hydrolysed to pump glucose across the membrane
- A clear channel is formed, connecting two sides of the membrane
- Glucose movement is facilitated by inward and outward facing conformations- alternating access
- Membrane potential is exploited to move glucose across the membrane
Where do primary active transporters get their energy from?
Use a primary source of energy, e.g ATP
Where do secondary active transporters get their energy from?
Two different substances are coupled. One molecule moves along its concentration gradient
How much quicker is passive transport than active transport?
10^4-10^8 times.
What does the jardetzky model postulate?
Transporter with alternating access- 1 side open at a time. The cavity which recognises the substrate has high affinity. When a conformational change takes place, binding affinity is reduced to release the substrate to the other side of the membrane.
This requires energy
Explain what lac permease is
Secondary active transporter which couples lactose to a proton gradient. ATP synthase maintains the proton gradient
How did the 12 transmembrane helices of lac permease arise?
12 transmembrane helices arisen from a duplication and fusion of a 6 transmembrane helix
Describe lac permease structure
12 transmembrane helices
Kinked helices for alternating access
Central hydrophilic cavity for sugar binding site
Two halves separated by a loop between TM6 and TM7