Membranes: Structure and Transport Flashcards
What are the functions of membranes?
Seperation of compartments to create different environments within one cell, to distinguish self from not self, to exclude toxic compounds and to keep metabolites.
Communication with the exterior via exchange of signals and transport in both directions
Energization via photosynthesis and respiration and the providing of an electrical barrier
Outline the structure of a phosphoglyceride molecule
Choline-Phosphate=Glycerol=2xFatty Acids
What are the two structures lipids form in water?
Conical => Micelle
Cylindrical => Lipid bilayer
This is dependent upon the ratio of hydrophillic head to hydrophobic tail
Why are liposomes formed?
A planar sheet of phospholipid bilayer has esges exposed to the water, this is energectically unfavourable therefore they curl up to form a sphere.
Roughly how thick is a cellular membrane’s inner space
5-7nm
What is FRAP?
Fluorescence Recovery after Photobleaching. An small spot of a stained cell membrane is bleached with a laser. After time the colour recovers. This is proof of the fluidity of membranes
What is the rate of lateral movement of phospholipids in a typical cell?
10^7 times per second
What is the fluidity of a membrane dependent upon?
How saturated the hydrocarbons are
Temperature
Cholesterol
What is the most dominant form of lipid in the cell membrane?
Glycerolipid
What are the different types of phosphoglycerolipids?
Phosphatidylserine
Phosphatidylcholine
Phosphatidylethanolamine
Phosphatiidylinositole
How is a glycerolipid synthesised?
L-Glycerol 3-phosphate has an acyl group and R chain added to it by acyltransferase to create Lysophosphatidate which has an acyl group and R1 group added to it by acyl transferase to create a phsophatidic acid.
What are the 4 types of sphingolipids?
Sphingosin
Ceramid
Sphingomyelin
Glycosyl-cerebroside
How is a sphingolipid synthesised?
Palmitoyl-CoA reacts with serine to form Beta-Ketosphinganine which NADPH reduces to Sphinganine which reacts with Fatty acyl CoA to form N-acylsphinganine, which then reacts with UDP-Glucose to become Cerebroside and Phosphatidylcholine to become Sphingomyelin.
What are the 3 glyosphingolipids?
Galactocerebroside
GM1 ganglioside
sialic acid
What are the three sterols?
Cholesterol
Egosterol (yeast)
Siststerol (plants)
What is a scramblase and what does it do?
Phospholipid synthesis only adds to the cytosolic half of the bilayer, scramblase catalyses the flipping of molecules to the outer leaflet so that the membrane grows symmetrically.
What are the typical percentages in the inner and outer leaflets?
Inner leaflet - Cholesterol 3%, Phosphatidyl choline 10%, Phosphatidylethanolamine 20%, Phosphatidylserine 15%, Phosphatidylinositole, Sphingomyeline,
Outer leaflets - Cholesterol 5%, Phosphatidylcholine 20%, Phosphatidylethanolamine 10%, Phosphatidylinositole, Sphingomyeline, Glycosphingolipids
What is one of the membrane associated markers of apoptosis?
Presence of phosphatidylserine on the outer leaflet
What does a flippase do?
They are responsible for catalysing flip-flopping across the membrane so that different phospholipids are on different leaflets.
How does a flippase work?
It doesn’t have enzymatic activity as such but rather encloses a hydrophillic moeity so that the phospholipid can flip across to the other leaflet
What is a phospholipase?
Hydrolyses phospholipids into fatty acids and other lipophillic substances
What does phospholipase A1 cleave?
SN-1 acyl chain
What does phospholipase A2 cleave?
SN-2 acyl chain, releasing arachidonic acid
What does phospholipase B cleave?
cleaves both SN-1 and Sn-2 acyl chains
What does phospholipase C cleave?
Cleaves before the phosphate releasing DAG and a phosphate containing head group
What does phospholipase D cleave?
Cleaves after the phosphate releasing phosphatidic acid and an alcohol
Which phospholipases are considered phosphodiesterases?
C and D
There are three shapes of lipids. What are they and which lipids tend to take this form?
Cylindrical - Most GPLs
Conical /\ - cholesterol
Conical - \/ - Sm, GSLs and some GPLs
What do the varying shapes of phospholipids suggest?
That they group together in microdomains to form lipid rafts
What causes alternating thickness in membranes?
Lateral segregation of saturated and unsaturated hydrocarbons
What experiment proves lateral segregation?
Artificial membrane vesicles with different stains so theblue is the lipid ordered state rich in sphingomyelin and cholesterol. The red is the disordered state rich in dioleoylphosphatidylcholine. This staining shows clustering dependent on the ratio of lipids.
What experiment shows the existence of membrane proteins?
When measuring the permeability of the lipid bilayer against the permeability of a biological membrane you find that it deviates for K, Na, Cl and h20 so that they are transported faster in a biological membrane suggesting that there sis ome form of protein which does this.
What is the percentage of genes that encodes membrane proteins?
30-40%
What is the sandwich model?
Proposed by Davson and Danielli, they proposed that proteins lined the outer side of the lipid bilayer like a sandwich. However this does not take into account the fact that in order to facilitate membrane transport the protein must span the hydrophobic portion if the membrane
How was it found that proteins are actually integral to the membrane?
Using freeze fracturing they found that the inside of the extracellular layer was smooth and the inside of the cytoplasmic layer was bumpy, thus disproving the sandwich model
What experiment shows that membrane proteins can move laterally?
The fusion of a tagged mouse cell, so that membrane proteins show up in a certain colour, and a tagged human cell to form a heterozygote by the sendai virus. After about 40 minutes the membrane proteins are evenly distibuted across the cell.
What is the fluid mosaic model?
Outlined by Singer and Nicholson it describes the membrane as a mosaic of proteins dispersed unevenly in the bilayer, with only their hydrophillic regions exposed to water.
What is the lipid raft concept?
Proposed by Simons and van Meer it says that proteins are clustered together dependent upon their function. This is currently under debate.
What are the problems with the lipid raft model?
A line tension should exist between the L alpha and Lo phases. This has been seen in model systems but not in cell systems.
There is no consensus on lipid raft size
The time scale of lipid raft existence is unkown. they may occur on a timescale irrelevant to biological processes.
What are the functions of membrane proteins?
Transport Enzymatic activity Signal transductionCell-cell recognition Intercellular joining Attachment to the cytoskeleton and ECM
What are the problems of structural analysis of membrane proteins?
They have a low abundance compared to cytosolic protein
Extraction from membranes requires detergent
tedious/impossible to get enough material
Expression bottleneck
Poor stability in solution
Poor crystallisation
Low resolution mebrane analysis techniques
Prediction of transmembrane alpha helices or beta barrels
topology = no. of Tm domains, orientation of termini
Validation by means of analyses of fusion proteins whose orientation is already detectable
What are the structural classes of membrane proteins?
bitopic Polytopic monotopic FA or Prenyl residue GPI-anchor Bound to an integral protien
What is the equation for the chemical potential of an uncharged substnace?
potential = RT ln Cis/Cos
give an example of a uniporter
GLUT1
What is the chemiosmotic theory?
Chemical energy is stored in concentration gradients of ions across membranes
How can heterologous expression be used to investigate transporters?
If you knock out the 3 endogenous ammonium transporters in yeast you can investigate how to reestablish transport.
What are the testable parameters and limitations when investigating cell membrane transport?
Complemenatation
biochemical parameters
inhibitors
But most data are a mix of parameters
What is a common model for studying transporters?
Xenopus laevis oocytes
