M&R Session 1: membrane proteins and cytoskeleton Flashcards
What are the 6 functions of biological membranes?
- Barrier
- Control
- Communication
- Recognition
- Signal generation
- Some specialised functions
In order of decreasing percentage composition, what substances is a membrane bilayer made up of?
Protein
Lipid
Water
Carbohydrate
Membrane lipids are amphipathic. Give some examples of lipids in membranes
Phospholipids-predominant lipid, range of polar head groups, usually C16-18, Cis double bond introduces a kink so decreased packing. Fatty acid + glycerol + phosphate head. FAs attached to C1&C2 of glycerol, PL groups can be choline/amines/AAs/sugars
Sphingomyelin-no glycerol backbone
Glycolipid a-fatty acid with carbohydrate
Name the two types of glycolipid structure found in membranes
Cerebroside: one sugar attached to head
Ganglioside: oligosaccharide head group
Functions of membrane proteins?
Enzymes, transporters, pumps, ion channels, energy transducers, structural elements
Which has more protein in its membrane: myelin or mitochondria?
Mitochondria
How are lipid bilayers formed and stabilised?
Favoured structure for PL and GL in aqueous media. Formation: van der Waals forces between hydrophobic tails
Stabilised: electrostatic and hydrogen bonding between hydrophilic heads, and interactions between the hydrophilic groups and water
How can lipids move in a membrane?
- Intra-chain motion: vibrations and kinks in FA chains
- Fast axial rotation
- Fast lateral diffusion within the plane of the bilayer
- Flip-flop: 1 for 1 exchange of lipids between two halves of the bilayer (limited)
How may integral proteins move in a membrane?
Conformational change
Rotational
Lateral diffusion
Why can membrane proteins not ‘flip-flop’?
Not thermodynamically favourable. As large hydrophobic groups would have to move through a hydrophobic core
What mobility restraints apply to membrane proteins?
Lipids-proteins tend to separate out into cholesterol-poor regions
Membrane protein associations e.g. with peripheral proteins on the cytoskeleton
Protein aggregates
Tethering
Interaction with other cells
Describe the structure of cholesterol
Rigid planar steroid ring structure, with a polar head group and a non-polar hydrocarbon tail
How does cholesterol regulate membrane fluidity?
Increases fluidity by reducing phospholipid packing due to its rigid planar ring structure
Decreases fluidity by decreasing phospholipid tail mobility as the rigid ring is held close to the FA chains
Describe two ways that a membrane protein may interact with the hydrophobic domain of a membrane bilayer
- Transmembrane sequence of 20-22 amino acids with hydrophobic R groups. Could be alpha helices or beta sheets
- Lipid-linked proteins through insertion of hydrophobic lipid tail, e.g. post-translational modification with a fatty acid
What is the evidence for the presence of proteins in membranes?
Functional: facilitated diffusion, specificity of cell responses, ion gradients. Biochemical: membrane fractionation and gel electrophoresis
Describe the two types of membrane protein
Peripheral: bound to surface by DSBs, electrostatic interactions and HBs, can be removed by changes in pH or ionic strength
Integral: interact with hydrophobic regions, cannot be removed by altering pH/ionic strength, removed by detergents or organic solvents that compete for the non-polar interactions
Which cell is used as a model plasma membrane?
Erythrocytes. Erythrocyte ghosts prepared by osmotic haemolysis to relese cytoplasmic components, these are analysed by gel electrophoresis and SDS-PAGE
How are proteins on erythrocyte membranes identified?
Peripheral: released after treatment with high ionic strength/changing pH, located on cytoplasmic face
Integral: only dissociate with detergent, protein bands 3 and 7
Both types are glycoproteins as they have covalent CHO groups
What is the purpose of CHO groups in erythrocyte membranes?
Very hydrophilic CHOs prevent flip flop
Specific CHOs for cellular recognition
How can proteins associate with a membrane?
Single or multiple transmembrane domains
Post translational lipid modifications e.g. mystroylation, palmitoylation
Sulfhydryl groups
Dolichol phosphate-linked polypeptide
Describe the structure of the cytoskeleton
Alpha and beta subunits of SPECTRIN (long floppy rod) wind together to form an ANTIPARALLEL HETERODIMER, then two of these join to form an A2B2 HETEROTETRAMER
Rods are crosslinked into networks by short ACTIN protofilaments, band 4.1 and adducin molecules (interact end of spectrin rods)
How does the cytoskeleton interact with the membrane?
Spectrin-actin network attached through adaptor proteins.
Peripheral membrane proteins-spectrin, actin, band 4.1, ankyrin (band 4.9), adducin
Integral membrane proteins: band 3 (anion exchanger), glycophorin A
What is hereditary spherocytosis and why does it lead to haemolytic anaemia?
Spectrin levels depleted by about 50%, erythrocyte cytoskeleton weakened, erythrocytes become spherical rather than biconcave, become trapped in capillaries, are lysed, cleared by spleen. Bone marrow can’t compensate for the decrease in RBCs so anaemia results
What is hereditary elliptocytosis?
Defect in spectrin meaning it can’t form heterotetramers, causing fragile elliptoid cells that lyse more easily
Describe the steps in membrane protein biosynthesis from recognition of signal sequence to ribosome detachment
- N-terminal signal sequence recognised by SRP, inhibiting protein synthesis while ribosome is in cytoplasm
- In ER, SRP recognised by receptor and signal sequence released to interact with the receptor, causing further synthesis through ER membrane
- Ribosome anchors to pore complex through which polypeptide chain is extruded
- Stop sequence reached, signal sequence cleaved from new protein by signal peptidases
Describe the new transmembrane protein orientation
N terminal in lumen, C terminal in cytoplasm
What happens to the transmembrane protein after it leaves the ER?
First fold is the driving force for other transmembrane domains. The nascent chain is processed further as it passes from cis to trans Golgi. Secreted proteins are released to membrane, so regions located in the cytoplasm during synthesis remain with this orientation
Describe post translational modifications of membrane proteins
Core CHO transferred from a dolichol phosphate carrier lipid
Protein disulfide isomerases catalyse formation of DSBs whilst protein is in the ER
Final CHO modifications in trans Golgi
Glycosylation is required if proteins are targeted to lysosomes
