Intro to Ion Channels Flashcards
What is membrane transport?
The selective movement and redistribution of solutes across biological membranes.
What are examples of membranes involved?
Plasma membranes.
Vacuoles.
Mitochondria.
Chloroplasts.
Endoplasmic reticulum (ER).
Sarcoplasmic reticulum (SR).
Lysosomes.
What is the importance of membrane transport?
Osmoregulation / turgor control.
Nutrient acquisition.
Waste excretion.
Compartmentalisation of metabolism.
Energy transduction.
Signal transduction.
What are the 3 classes of transport system?
Primary pumps.
Carriers.
Ion channels.
How do primary pumps work?
Use a primary source of energy to pump ions against electrochemical gradients
Primary active transport
Ions = usually H+ or Na+
Pumps are electrogenic.
Establish electrochemical gradients for ‘driver’ ions.
How do carriers work? (general)
Energised by ‘driver’ ion electrochemical gradients
Secondary active transport.
What are the classes of carriers?
Symporters / co-transporters
= move solute against gradient
Antiporters / counter-transporters
= get rid of waste solute in opposite direction
Facilitators / uniporters
= cannot be energised, allow facilitated diffusion passively down electrochemical / chemical gradients
How do ion channels work?
Transport is always passive down electrochemical gradients.
Usually highly regulated with defined open and shut kinetics = gating.
Usually selective for specific ions.
What are the 3 distinct components of signalling networks?
Perception of signal.
Intracellular signalling.
Cellular responses.
= ion channels involved in all 3
What are the 4 basic properties of ion channels?
Permeation.
Selectivity.
Gating.
Modulation.
What is permeation?
When a channel opens = permeant ions move through.
Direction governed by electrochemical gradient.
Rate governed by electrochemical gradient / intrinsic properties of channel pore.
What is selectivity?
Simple = distinction between cations and anions.
Other channels more complex = can distinguish between size
Non-selective = permeable to anions and cations.
What is gating?
The process of transition between the open and closed states is gating.
What is the frequency or opening / closing directly influenced by?
Changes in membrane voltage (voltage gated).
Binding of intracellular or extracellular ligands (ligand gated).
Mechanical stress (mechano-sensitive).
Changes in temperature.
Which substances can modulate gating of channels?
Ca2+, H+, ATP, Fatty Acids, Phosphorylation, G proteins.
= they influence gating following activation (indirectly)
What are accessory proteins?
Usually present the pore forming unit.
Most channel possess tightly associated accessory subunits.
What are some functions of accessory proteins?
Specifying their location and abundance of ion channel
= Rapsyn clusters AChR at excitatory synapses
Modulate ion channel gating
= SUR subunit of KATP channels has key role in hydrolyzing ATP to stimulate channel opening
Fine tuning sensitivity to physiological and pharmacological agents
= SUR is target of drugs used to treat diabetes
How to study ion channels?
Electrophysiology
= properties / activity
Cloning and determining primary sequence
= physiological role
Topology
= structure / function
How to clone ion channels?
Size fractionate on a gel to form different pools of mRNA and inject into oocytes (e.g. from Frog)
Electrophysiology.
Generate cDNA library (used to generate large amounts of mRNA).
Repeat fractionation and expression to isolate single mRNA.
Sequence mRNA - gene identified
e.g. anoctamins
What is another example of expression cloning?
Heterologous expression and complementation of yeast mutant growth phenotype
e.g. trk1D, trk2D mutant
What are the 6 principles of hydropathy analysis? (Topology)
(1) Assume hydrophobic polypetides in a hydrophobic environment (membrane) form apha helices.
(2) Membrane span is 6nm (equivalent to 20 a.a. in a helix)
(3) Each a.a. is given hydropathy index (reflects hydrophic/hydrophilic nature)
(4) Stretches of a.a. sequence are analysed by computer using a moving segment approach (20 a.a.)
(5) Hydropathy index is plotted
(6) Values over +1 are scored as forming transmembrane spans
What are the ion channel families?
Pore-loop channels.
S4 family.
Non-pore loop channels.
What are the 3 pore-loop families?
Kir family
= K+ inward rectifiers
= e.g. ATP sensitive, G-protein coupled
= K+ influx, regulate membrane voltage
Two pore K+ channel family
= tandem duplicate of Kir
= 15 members
= represent ‘leak’ channels
Glutamate receptor family
= all cation permeable
= pore loop in inverse direction
= extracellular binding domain
What are examples of the S4 family?
Voltage gated family
= S4 voltage sensor (depolarisation)
= Kv subfamily = K+ efflux, repolarisation of AP
= NaV subfamily = Na+ INFLUX, AP depolarisation
= CaV subfamily = Ca2+ signalling
SK + BK families
= Ca2+ activated K+ channels (efflux), neuron repolarisation
= SK subfamily = small G , voltage insensitive, activated in nM range of Ca2+
= BK subfamily = large G , activated by depolarisation and uM range of Ca2+, 7TMS
CNGC family
= photo and olfaction reception
= activated by direct binding to CNs, modulated by voltage
HCN family
= activated by hyperpolarisation, CN modulated
= identified in cardiac pacemaker cells
TRP family
=promiscuous activation
= voltage insensitive
= roles in sensory perception
What are the non pore-loop families?
Cys-loop receptor family
= pentamers, pore formed by TMS2
= bind neurotransmitter extracellular (N-terminus)
= large family, members according to agonist, synaptic transmission
= e.g. AChR, GABA,GlyR, 5HT
IP3 and RYR receptor families
= major intracellular Ca2+ release channels
= endomembrane location, large structures
= IP3 - a2+ signalling roles
= RYR - muscle contraction
CLC family
= anion selective
= complex structure / function relationships - channels and transporters
= located in plasma membrane (CLC1) and endomembrane (CLC5)
CFTR (ABC transporter)
= ABC family
= ATP hydrolysis used to drive protein between open and closed
= epithelial transport in humans
= e.g. cystic fibrosis
Bestrophin family
= anion selective, Ca2+ activated
= likely to form tetramers (TMS2 forms pore)
= possible PM / endomembrane location
= physiological function unclear
e.g. eye diseases - BEST disease (macular degeneration)
Anoctamines (TMEM16)
= Cl- selective (Ca2+ activated), 10 family members, 8 TMS
= thought to be ubiquitous cells found in all animal cells
Ion Channels are complex because they exibit:
Selectivity amongst ions.
Tight control over opening and closing = gating
Fine control or adjustment of gating = modulation
Complex structures (composed of 100s a.a)
Accessory proteins
Wide variety of structures, loosely arranged into distinct families based on secondary structure
Complex structure / function relationships
