Topic 3 Flashcards
(89 cards)
Primary protein structure
sequence of amino acids
Secondary structure
alpha and beta sheets as a result of the nature of the amino acid sequences
- Polar side chains will face the water
- Non-polar will wrap themselves inside of the protein structure
Tertiary structure
full length of the protein all folded up together in a 3D structure
Quaternary structure
subunits of proteins are bound together with other proteins to form a functional units
Domain
a certain region of a protein which can be structurally and/or functionally different
- Alpha helix structures like to be embedded in membranes which is good for ion channels
replication
DNA synthesis
Transcription
RNA synthesis
Translation
protein synthesis
what drives the folding of proteins?
the nature of the amino acids
alpha helix
ribbon like structure forming a helical structure
- Individuals proteins can have multiple helices and other types of secondary structures
- the charges are lined up and there are 0.54 nm between twists
Beta sheets
flat folds of amino acids that stack on top of one another and have 2-3 bridging H-bonds
- form a twisted, pleated sheet
what type of protein folding makes good transmembrane domains?
alpha helices
- the external region is hydrophobic which allows it to integrate into the membrane and the inside is hydrophilic which allows aqueous solution to flow through
- When they all come together they make a functional transmembrane protein used as a channel ⇒ ion flow
what are the ways proteins interact with the lipid bilayer? (4)
- transmembrane
- mono layered
- lipid linked
- protein attached (to another protein)
what did early voltage clamp experiments tell us?
how current changes over time at a particular voltage
- Required large axons to accommodate the relatively large electrodes
- These were whole cell currents ⇒ for the whole population of channels on the cell
what is different about the modern voltage clamp?
has much smaller/thinner electrodes
- Can record whole cell currents from single mammalian neurons
- Can record current through a single channel
Microscopic current
any current that flows through an individual ion channel
Patch clamp
glass needle microelectrode pulled to a fine and sharp point attached to an amplifier ⇒ touches wherever you want to record in the cell
- The electrode has an internal core thats hollow that we can fill up with saline solution and when we apply a mild suction it will pull the membrane to make a tight seal against the glass
- Current cannot flow in our out of the electrode ⇒ you can get a single ion channel in the tip of the electrode
how are the patch clamp and voltage clamp similar?
the patch clamp is a subtle version of voltage clamp
T/F It is possible to clamp voltage and current
False
- not both at the same time
Whole cell
give it a strong transient suction which pops a hole in the membrane so the cytoplasm is continuous with your recording electrode
Single channel inside out recording
move the electrode away from the cell out of the aqueous environment into air and then back in so now you have an individual ion channel that you can record for current
how do patch clamps work in frog eggs?
- Get a frog egg and inject DNA
- Overexpression of the protein will be expressed and you get lots of channels in the membrane of the egg
- You can use a patch clamp recording device to use whole cell or inside out recording
how does using a patch clamp with TEA show data?
Blocks the K+ channels and has a transient depolarization causing inward current that disappears right away
- the K+ channels inactivate and stay inactivated during the rest of the pulse
- some Na+ channels chatter on and off but have a delayed start
- on a macroscopic scale the cell has an inward current which dips the graph down before it rises back to current 0 and stays there
how does using a patch clamp with TTX show data?
Depolarization pulse causing an outward current that stays longer than the Na+ inward current before dropping back down
- the K+ channels will open a little bit later but mostly stay on causing an increase in the microscopic measurements and macroscopic measurements
- once a channel opens it will stay open