Principles of Cell Biology (BIOL2060)

Department of Biology
Memorial University of Newfoundland

Membranes: Structure, Function and Chemistry

Functions of Cell Membranes
1) Define boundaries (permeability barrier)
2) Sites of specific functions
3) Regulation of solute transport
4) Signal detection and transmission
5) Cell to cell communication

Structure of Cell Membranes
1) Lipids as a bilayer
2) Proteins may transverse the lipid bilayer via alpha helices; integral proteins

Models of Membrane Structure

"The Fluid Mosaic Model" (Singer & Nicholson: 1972)
Proteins are retained the lipid bilayer and proteins may also be discontinuously embedded in the membrane: integral proteins

Integral proteins have one or more series of hydrophobic sequences that span the lipid bilayer (Unwin & Henderson 1975)
i.e. bacteriorhodopsin: a single peptide chain folded back and forth across the lipid bilayer 7 times!

The Fluid Mosaic Model is universally accepted model for cell membrane structure.
"Mosaic of proteins in a fluid lipid bilayer"
1) Lipids are fluid and this fluidity can be regulated.
2) Proteins form a "mosaic", perform a variety of functions, and protein fluidity may be restricted.
3) Both are asymmetrically distributed in membranes.
 

Membrane Lipids and the Fluid Part of the Model

Membrane Lipids: Classes
Three classes of membrane lipids
1) phospholipids - most prominent component
2) glycolipids - addition of carbohydrate/sugar group
Lipids are amphipathic molecules that have
hydrophobic or fatty-acids tails
and a hydophillic or polar head

The fatty acids tails can be of differing length and (saturation or) number of double bonds (12 - 20 carbon atoms long)
3) sterols - rings in their structure
hydrophobic
e.g. cholesterol, ergosterol, and phytosterols
(these absent from prokaryotes & inner membrane of mitochondria and chloroplasts).

Membrane Lipids: Behaviour

Asymmetry
Little or no transverse diffusion (flip-flopping) from one lipid layer to the other
e.g. glycolipids - outside
e.g. phosphatidlyethanolamine ? inside
Note that enzymes known as flippases exist that under special conditions do this.

Fluidity
1) rotational movement
2) lateral diffusion within the lipid layer
demonstrated experimentally by fluorescence recovery after photobleaching
(a few microns per second or less)
Functioning of the lipid bilayer and thus the membrane is affected by temperature.
Transition temperature (Tm): temperature at which a membrane changes between the fluid and gelled state
The Tm is effected by
1) the length of the fatty acids
2) the number of double bonds &
3) the proportion of sterols (e.g. cholesterol)

Tm increases as the length of the fatty acid increases

Tm decreases as the number of double bonds increases

Sterols (e.g. cholesterol) moderate in both directions!
Acts as a buffer
up to 50% in mammalian cells
Less fluid at higher temps
More fluid at lower temps

Organisms regulate membrane fluidity primarily by changing lipid composition
cooler - shorter fatty acid chains
cooler -  increase the number of double bonds

Poikilotherms & homeoviscous adaptation
e.g. Micrococcus - enzyme cuts two carbons off the 18 carbon fatty acids to produce 16 carbon fatty acids in cool temperatures

E. coli - desaturase enzyme increases number of double bonds in cool temperatures

cold-hardy plants: desaturase enzyme activity triggered at lower temperatures

warm blooded hibernating mammals:
increase numbers of double bonds

Membrane Proteins and the Mosaic Part of the Model

Membrane Proteins

Freeze fracture microscopy shows us that proteins are embedded or "floating" in and on the lipid bilayer of cell membranes

Correlates well with the known protein content of cell membranes
e.g. erythrocyte (red blood cell: RBC) membrane  vs the inner mitochondrial membrane

Proteins, like lipids, show membrane asymmetry (freeze fracture microscopy)
many are glycosylated (2 - 60 sugar units!) = glycoproteins
The gycocalyx of animal cells for recognition in antibody binding and tissue formation thus membranes are often functionally asymmetric

Membrane Proteins: Main Classes
1) integral membrane proteins : monotopic or transmembrane
transmembrane proteins cannot be readily removed, require detergents
2) peripheral membrane proteins : weak electrostatic forces
3) lipid anchored proteins:  covalently bound

Transmembrane proteins have one or more transmembrane segments
cannot be readily removed require detergent to isolate

Membrane Proteins: mobility
1) some move freely
2) others are anchored:
e.g. on the inside to the cytoskeleton (scale?)
e.g. on the outside to the ECM
e.g. integrin - cell attachment and signalling
Or via lipid rafts

Some move freely as shown by fluorescent antibody tagging and cell fusion
or by putting the cell in an electrical field

Membrane Proteins: Methods
Application of molecular techniques to the study of membrane proteins

X-ray crystallography vs hydrophobicity plots
Difficult for hydrophobic proteins but 1988 Nobel Prize for resolution of alpha-helical domains.

Proteins and SDS-polyacrylamide gel electrophoresis
Membrane proteins and Molecular Biology

Proteins can be separated by SDS-polyacrylamide gel electrophoresis = SDS-PAGE
Electrophoresis of Sodium dodecyl sulphate (SDS) coated proteins:
Coats with a negative charge which disrupts membranes and denatures proteins

May use the gel to identify specific molecules by immunoblot  (Western)
 

Transport Across Membranes

Cell and Transport Processes

The hydrophobic inner portion of the lipid bilayer allows some substances to pass through it.
However, it is an absolutely important barrier to a majority of other substances.

This permeability must be overcome to allow some important small molecules and ions, into and out of the cell.
 

How do substances cross membranes?

1) Simple diffusion ? via lipids
no cellular energy expended
Down the gradient toward equilibrium

2) Facilitated diffusion - protein mediated (carrier and/or channel proteins)
no cellular energy expended
Down the gradient toward equilibrium

3) Active Transport  - protein mediated
cellular energy expended
Up the gradient
 

Simple Diffusion

Occurs via lipid interaction; lipophilic/hydrophobic substances
Simple non-polar molecules
e.g. di-oxygen, carbon dioxide, carbon monoxide
e.g. some drugs (nicotine, heroin),  anesthetics,
The cell membranes are not (cannot be) selective in simple diffusion.

Facillitated Diffusion

Occurs via proteins: lipophobic/hydrophillic
Very dependant upon the proteins present in the cell membrane
This is a very specific process.
The cell membranes are very selective in facilliated diffusion of certain solutes.

A. carrier proteins
(= transporters, permeases)

B. ion channels
e.g.  red blood cell (RBC) glucose transporter
an integral membrane protein with 12 transmembrane segments
Glucose in blood plasma is about 3.6-5.0 mM but inside RBC it is about 0.5-1.0 mM  (=concentration gradient)
& because there is a glucose selective transporter in the RBC membrane

Facilitated diffusion occurs
50,000 X as fast as in a lipid bilayer alone
In this case, very selective for glucose and a small group of similar sugars

RBC anion exchange protein - antiport carrier protein
chloride and bicarbonate exchanger
reciprocal exchange
obligatory and very selective
used in removing carbon dioxide from the tissues to the lungs

Channel proteins
These proteins form hydrophilic transmembrane channels across the membrane
e.g . Ion channels are specific for one of the following: calcium ions, potasium ions, sodium ions, chloride ions
moves 1 million ions per second!
Regulated by changes in voltage, ligand-binding, and mechanical forces (i.e. stretching)
e.g. Nerve cells with BEST.

How do substances move across membranes against the concentration gradient?

Active Transport
Occurs via proteins and requires energy!
Active Transport is endergonic; and produces a thermodynamically unfavorable difference in concentrations.
Active Transport enables a cell to maintain a constant intracellular non-equilibrium concentration of specific ions etc.
e.g. Sodium Potassium Pump :
Direct active transport maintains electrochemical ion gradients in mammalian cells
maintains
potassium ions at a ratio of 35 inside: 1 outside   (35:1)
sodium ions at a ratio of 0.08 inside: 1 outside   (0.08:1)
Three sodium ions are moved out; 2 potassium ions are moved in for each ATP molecule hydrolyzed

This forms a electrochemical gradient
P-type: phosporylation event

Model of Mechanism of Na+/K+Pump
Functions of the Na+/K+ pump/ATPase:
maintaining osmotic equilibrium
maintains voltage difference across the membrane - conduction of electrical impulses by nerve and muscle cells
driving force for cotransport / indirect active transport

There are many different types pumps/ATPases:
P-type pumps
"phosphorylation"
eg. Na+ /K+ pump of animal cells, PM
eg. H+ pump of fungi and plants,
eg. H+ pump of the gastric epithelium;
eg. Ca++ pump of the sarcoplasmic reticulum
vanadate inhibition
ABC type pumps
ATP binding cassette
prokaryotes mainly
variety of solutes
can  pump antibiotics out of the cell: Antibiotic resistance

Multiple Drug Resistant Transport protein in tumour cells
Active Transport: Direct and Indirect
Indirect Active Transport
cotransport
one solute down its gradient
eg Na+ in animals, H+ in plants and fungi
which drives a second solute up its gradient
eg. monosaccharide or amino acid
symport or antiport
e.g. Sodium/Glucose Symporter
driven by sodium gradient
in epithelium of the intestine

Bacteriorhodopsin Proton Pump of Haloabacteria
Photon of light drives the Halobacterium (purple bacteria) proton pump.

Notes prepared from Becker's World of the Cell, 8th & 9th editions
Hardin, Bertoni & Kleinsmith, 2012, 2016
Figures copyright of Pearson Education Inc.
email me at bestave@mun.ca