Medical Biochem Week 10 - Biological Membranes

Biological Membranes

Function of Biological Membranes

• Define external boundaries of cells.
• Define internal boundaries of cells, such as organelles.
• Create environments inside cells that differ from the outside.
• Protect cells from their surroundings.
• Facilitate communication with other cells.
• Enable contact with other cells and the microenvironment.
• Selectively allow molecules to enter and exit cells.
• Fuse with other membranes.
• Enable cell division without leakage of contents.

Lipid Aggregation in Water

• Lipids aggregate into three major structures in water:
  • Micelles
  • Bilayers
  • Vesicles (liposomes)
• The structure formed depends on the type of lipid and its concentration.

Micelles

• Form in solutions of amphipathic molecules with larger, more polar heads than tails.
• Composed of a few dozen to a few thousand lipid molecules.
• Aggregation is concentration-dependent.
• Virtually no water in the interior.

Membrane Bilayers

• Form when lipids with polar head groups and more than one lipid tail are in an aqueous solution, such as phospholipids and sphingolipids.
• Hydrophilic head groups interact with water on both sides.
• Hydrophobic fatty acid tails are packed inside.

Vesicles (Liposomes)

• Small bilayers spontaneously seal into spherical vesicles in a concentration-dependent manner.
• Fuse readily with cell membranes or each other.
• Synthetic vesicles can be made with artificially inserted proteins.
• The central aqueous cavity can enclose dissolved molecules, such as drugs, for delivery.

Properties of Biological Membranes

Common Features of Membranes

• The main structure is a lipid bilayer.
• The lipid bilayer forms spontaneously in aqueous solution and is stabilized by noncovalent forces, especially the hydrophobic effect.
• Sheet-like flexible structure, $30-100 \AA$ (3-10 nm) thick.
• All cells have a cell membrane that separates them from their surroundings.
• Eukaryotic cells have internal membranes dividing the internal space into compartments (organelles).

Membrane Bilayer Population

• Proteins are embedded in or associated with the membrane.
• Integral proteins are firmly associated, often spanning the bilayer.
• Peripheral proteins are weakly associated and easily removed.
  • Some are noncovalently attached.
  • Some are linked to membrane lipids.

Fluid Mosaic Model of Biological Membranes

• Membrane proteins and lipids can rapidly diffuse laterally or rotate within the bilayer; proteins "float" in a lipid-bilayer sea.
• Uncatalyzed lateral diffusion: Individual lipids undergo fast lateral diffusion within the leaflet.
• Uncatalyzed transbilayer (flip-flop) diffusion between the leaflet is rare because it is difficult for the charged head group to transverse across the hydrophobic tail region.
• Special enzymes catalyze transverse diffusion.
• Flippases are unique unidirectional and bidirectional enzymes that catalyze lipid movement.
• Some flippases use the energy of ATP to move lipids against the concentration gradient.
• PE: phosphatidylethanolamine
• PS: phosphatidylserine

Methods to Illustrate the Fluid Mosaic Model

• Measurement of lateral diffusion rates of lipids by fluorescence recovery after photobleaching (FRAP).
  1. Label lipids in the outer leaflet of the plasma membrane with a fluorescent probe.
  2. Bleach a small area with an intense laser beam.
  3. With time, labeled lipid molecules diffuse into the bleached region, and it becomes fluorescent again.

Membrane Composition

• Lipid composition varies by:
  • Organisms
  • Tissues
  • Organelles
    • Cholesterol is predominant in the plasma membrane but virtually absent in mitochondria.
• Membranes are approximately 25-50% lipid and 50-75% protein.
• Lipids include glycerophospholipids, sphingolipids, and cholesterol (in some eukaryotes).
• The ratio of lipid to protein varies.
• The type of phospholipid varies.
• The abundance and type of sterols vary.
  * Prokaryotes lack sterols.

Membrane Proteins

Types of Membrane Proteins

• Integral Proteins: Firmly attached to the membrane and can only be removed by detergents.
• Peripheral Proteins: Loosely associated with the membrane and can regulate membrane-bound enzymes or limit the mobility of integral proteins.
• Amphitropic Proteins: Found in both the cytosol and associated with the membrane.

Six Types of Integral Membrane Proteins

• Classified based on spatial arrangements of protein domains relative to the lipid bilayer:
  • Type I & II: 1 transmembrane (TM) helix
  • Type III: Multiple TM helices from the same polypeptide
  • Type IV: Multiple TM helices from multiple polypeptides forming a channel
  • Type V: Covalently linked lipids
  • Type VI: TM and Glycosylphosphatidylinositol (GPI) anchors

Functions of Proteins in Membranes

• Receptors: Detecting signals from outside.
  • Light (opsin)
  • Hormones (insulin receptor)
  • Neurotransmitters (acetylcholine receptor)
  • Pheromones (taste and smell receptors)
• Channels, gates, and pumps:
  • Nutrients (maltoporin)
  • Ions (K-channel)
  • Neurotransmitters (serotonin reuptake protein)
• Enzymes:
  • Lipid biosynthesis (some acyltransferases)
  • ATP synthesis ($F<em>0F</em>1$ ATPase/ATP synthase)

Membrane Fusion

• The ability to fuse with another membrane without losing continuity.
• The 'fingerprint' of a membrane prevents or allows fusion of certain types of membranes.
• Many functions for this mechanism are found in eukaryotic cells.
• Membranes must:
  1. Recognize each other.
  2. Appose surfaces (get in close proximity).
  3. Disrupt bilayers.
  4. Fuse bilayers.
• Membrane fusion during Neurotransmitter release at synapse.
  • SNAREs and SNAP25 are targets of powerful neurotoxins.
  • Examples: Clostridium botulinum toxin, Tetanus toxin, Clostridium tetani.

Pores and Channels

• Pores and channels are transmembrane proteins with a central passage for ions and small molecules.
• Solutes of appropriate size, charge, and molecular structure can diffuse down a concentration gradient.
• Passive transport requires no energy.
• Active transport requires energy.
• The central passage allows molecules and ions of certain size, charge, and geometry to transverse the membrane.

Passive Transport

• Does not require an energy source.
• Protein binds solutes and transports them down a concentration gradient.
• The rate of transport is proportional to the concentration gradient of molecules across the membrane.
• Can be simple or facilitated.
• Uniport: A transporter carries only a single type of solute.
• Co-transport: Two solutes are transported.
  • Symport: Solutes move in the same direction.
  • Antiport: Solutes move in opposite directions.

Kinetics of Passive Transport

• The initial rate of transport increases until a maximum is reached (the site is saturated).

Model for Glucose Transport into Erythrocytes by GLUT1

• Passive transport
• Two conformational forms:
  • T1: Binding site exposed on the outside surface.
  • T2: Binding site exposed on the inner surface.
  1. Glucose in the blood binds to a stereospecific site and lowers the activation energy.
  2. A conformational change affects the transmembrane passage of glucose.
  3. Glucose is released into the cytoplasm.
  4. The transporter returns to the T1 conformation to transport again.

Active Transport

• Requires energy to move a solute up (against) its concentration gradient.
• The transport of charged molecules or ions may result in a charge gradient across the membrane.
• Primary active transport.
• Secondary active transport.

Primary Active Transport

• Powered by a direct source of energy, such as ATP, light, or electron transport.
• A solute is transported against its concentration gradient.

Secondary Active Transport

• An ion (S1 – often $Na^+$) is moved by primary active transport (requires ATP).
• This provides energy to drive co-transport of a second solute (S2) against its concentration gradient.
• A symport system (two solutes in the same direction) is used, but can also have antiport active transport.

General Mechanism for both Passive and Active Transport

• Protein binds a specific substrate.
• A conformational change allows the molecule or ion to be released on the other side of the membrane.

Example of Secondary Active Transport in Animals

• $Na^+-K^+$ ATPase of the plasma membrane.
• Responsible for maintaining low [$Na^+$] and high [$K^+$] in the cytosol.
• ATP is used to transport $Na^+$ against the concentration gradient.
• Leads to a net separation of charge across the membrane.
• Membrane potential ranges from -50 to -70 mV.
• The membrane potential then drives the import of solutes such as glucose.

Glucose Transporters in the Human Genome

• A table is provided listing various glucose transporters, their tissue expression, genes, and roles.
• GLUT1: Ubiquitous; basal glucose uptake.
• GLUT2: Liver, pancreatic islets, intestine; removes excess glucose from blood in the liver, regulates insulin release in the pancreas.
• GLUT3: Brain (neuronal); basal glucose uptake.
• GLUT4: Muscle, fat, heart; activity increased by insulin.
• GLUT5: Intestine, testis, kidney, sperm; primarily fructose transport.

Glucose Transporter 4 (GLUT4)

• Ingestion of carbohydrate-rich meals results in increased blood glucose.
• Excess glucose is taken up by cardiac and skeletal muscle (stored as glycogen) and adipocytes (stored as triacylglycerols).
• Regulated by the GLUT4 transporter and driven in response to insulin.
• Between meals, some GLUT4 are on the membrane, but most exist as intracellular vesicles.

Diseases of Ion Channels

Diseases of Chloride Channels

• Many diseases have been discovered by observing mutations (polymorphisms) in genes that code for ion channels.
• Chloride channels have various important functions:
  • Regulation of pH
  • Solute transport
  • Cell migration, proliferation, and differentiation
• Improper functioning of chloride channels = disease

Examples of Chloride Channel Diseases

• Dent’s disease: Inherited rare X-linked recessive condition affecting kidneys. Mutation in gene CLCN5 that codes a kidney-specific voltage-gated chloride channel. Most common cause of kidney stones.
• Thomsen disease: Dominant muscular genetic disorder characterized by muscle stiffness and inability to relax after voluntary movement. Affected muscle functions normally after a few repetitions.
• Becker disease: X-linked recessive inherited. A type of muscular dystrophy. Slowly progressive muscle weakness in legs and pelvis. Less severe than other MD types.

Diseases of Potassium Channels

• Long QT syndrome: Some are inherited life-threatening defects in the heartbeat (abnormal rhythm).
• A form of epilepsy, a rare inherited tendency to epileptic seizures in the newborn (abnormal neural integration). Affects $K^+$ channels in the brain, KCNQ2 and KCNQ3.
• Jervell and Lange-Nielsen syndrome: Several types of inherited deafness (affects $K^+$ gates expressed in the inner ear).

Diseases of Sodium Channels

• Some present as inherited disorders leading to certain types of muscle spasms.
• Cardiac disorders, some types of epilepsy.
• Liddle’s Syndrome: A single mutant allele yields a mutated channel. Too much $Na^+$ is reabsorbed and too little is excreted in the kidney.
• Leads to elevated osmotic pressure of the blood and results in hypertension.

Table of Diseases Resulting from Ion Channel Defects

• A table summarizes diseases, affected genes, and corresponding ion channels:
  • $Na^+$ (voltage-gated, skeletal muscle): SCN4A, Hyperkalemic periodic paralysis (or paramyotonia congenita)
  • $Na^+$ (voltage-gated, neuronal): SCN1A, Generalized epilepsy with febrile seizures
  • $Na^+$ (voltage-gated, cardiac muscle): SCN5A, Long QT syndrome 3
  • $Ca^{2+}$ (neuronal): CACNA1A, Familial hemiplegic migraine
  • $Ca^{2+}$ (voltage-gated, retina): CACNA1F, Congenital stationary night blindness
  • $Ca^{2+}$ (polycystin-1): PKD1, Polycystic kidney disease
  • $K^+$ (neuronal): KCNQ4, Dominant deafness
  • $K^+$ (voltage-gated, neuronal): KCNQ2, Benign familial neonatal convulsions
  • Nonspecific cation (CGMP-gated, retinal): CNCG1, Retinitis pigmentosa
  • Acetylcholine receptor (skeletal muscle): CHRNA1, Congenital myasthenic syndrome
  • $Cl^-$: CFTR, Cystic fibrosis