M&R 1.2 Membrane Proteins Flashcards
Describe the 2 types of evidence for proteins being present in membranes
- Functional (facilitated diffusion, ion gradients, specificity of cell responses)
- Biochemical (gel electrophoresis, freeze fracture)
How does freeze fracture work?
Lipid bilayer is frozen in ice, then fractured with a knife
The ice crystal will break at the weakest point (between the lamellae of the bilayer)
Proteins will be attached to one lamella and on the other one there will be a corresponding hole where the protein used to be
Describe the 3 ways that PROTEINS can move within bilayers
- Conformational change (e.g. to move things from inside to outside)
- Rotational (movement around its own axis)
- Lateral (e.g. to find a partner - dimerisation of receptors)
What way can proteins NEVER move in bilayers, and why?
Flip-flop
Because it’s so energetically unfavourable - the large hydrophilic moieties would have to pass through the hydrophobic middle of the bilayer in order to switch sides (would require loads of energy!)
What are 3 restrictions on protein mobility in membranes?
- Lipid- mediated effects (in general, proteins tend to be found in lower cholesterol, more fluid regions)
- Membrane protein associations
- with each other (e.g. at synapses)
- with proteins from other cells
- Association with extra-membranous proteins (tethering - e.g. to the BM or to the cytoskeleton)
How are peripheral membrane proteins associated with the bilayer?
They are associated with the membrane, but not within it
Bound to surface by electrostatic & H-bond interactions
How can peripheral proteins be removed from a bilayer?
They can be ‘washed off’ by changes in pH or charge
How are integral proteins associated with the bilayer?
They are transmembranous - they interact extensively with the hydrophobic domain of the bilayer
Can integral proteins be removed by changing pH or charge? If not, how can they be removed?
No
They can be removed by detergents and organic solvents (compete for non-polar interactions and essentially dissolve the bilayer to remove the embedded proteins)
What is the fluid-mosaic model of membrane structure? (Singer-Nicholson model)
Membrane proteins float in a ‘sea’ of lipid
Fluid - because unattached components of the membrane can move around freely
Mosaic - because made up of a patchwork of molecules
Transmembrane spanning domains are often what secondary structure?
Alpha helix
Transmembrane domains are usually ________ amino acids long
18-22 AAs long
Name some AAs that may be found in a transmembrane domain
AAs with hydrophobic R-chains: cysteine, leucine, alanine, phenylalanine
If you know the amino acid sequence of a protein but you don’t know where its transmembrane domains are, what would you use to find this out?
A hydropathy plot
- this maps to find hydrophobic regions of ~18-22AA’s that are likely to be TM domains
Aside from having single/multiple TM domains, describe some other different ways in which proteins can interact with bilayers
- Post-translational lipid modifications (proteins have bits of lipid/FA attached onto their extrinsic area, which lock it into the membrane)
- Dolichol phosphate-linked peptides (protein attached to dolichol phosphate (a carbohydrate) which is in turn attached to a lipid interacting with the membrane)
- Peripheral protein associations (peripheral proteins attached via association with integral transmembrane proteins)
How does membrane protein biosynthesis differ from secreted protein biosynthesis?
In membrane protein biosynthesis, there is a Stop Transfer Signal of 18-22 hydrophobic AA’s. When the ribosome translates it, this area of protein becomes stuck in the membrane because of it’s hydrophobic nature.
The ribosome gets pushed away as it translates the rest of the protein, which remains in the cytoplasm.
How are multiple transmembrane domains inserted in the membrane during membrane protein synthesis?
2 at a time (as pairs)
The rest build up as the protein is synthesised
Name 3 membrane proteins from an erythrocyte, and whether they are peripheral or integral
Spectrin - peripheral
Glycophorin (band 7) - integral
Band 3 protein - integral
The erythrocyte cytoskeleton is a network of ________ and ________ molecules, linked together by ________ and _________ molecules.
Network of spectrin and actin molecules, linked together by band 4.1 and adducin molecules.
In erythrocytes, the spectrin-actin network is attached to 2 different integral membrane proteins (band 3 & glycophorin A) by which adaptor proteins?
Ankyrin - attaches spectrin to band 3 protein
Band 4.1 protein - attaches spectrin to glycophorin A
Defects in the erythrocyte cytoskeleton can lead to which group of conditions?
Haemolytic anaemias (because the erythrocyte cytoskeleton is important for maintaining deformability)
How is the erythrocyte cytoskeleton affected in hereditary spherocytosis?
Spectrin levels are depleted by 40-50%
This makes the cage-structure more disparate - so RBC’s round up into spheres
How is the erythrocyte cytoskeleton affected in hereditary elliptocytosis?
There is a defect in the spectrin molecule
So even though there is plenty of spectrin, it cannot assemble properly (into heterotetramers)
Leads to fragile elliptoid (rugby ball-shaped) cells
How do hereditary spherocytosis/elliptocytosis lead to haemolytic anaemia?
The abnormally shaped cells are less resistant to lysis when passing through small capillaries
Therefore the cells break apart and get cleared by the spleen
Decreased RBC survival and bone marrow can’t compensate, leading to haemolytic anaemia
