ATP Powered Pumps Flashcards
What is active transport?
An endergonic process coupled to ATP hydrolysis, where a substance in low concentration is moved to an area of higher concentration on the other side of the membrane recquiring specific proteins
AKA primary active transport - the pumps use ATP as an energy source
Examples of molecules - ions, small hydrophilic molecules, lipids
What are the types of ion pumps used in active transport?
P-type ATPases - undergo phosphorylation as they transport cations
F-type ATPases - proton transporting complexes in mitochondria and bacterial membranes
V-type ATPases - proton transporting complexes in plant vacuoles and acidic vesicles (lysosomes)
ABC transporters -transport a variety of substances, including drug molecules
A-type ATPases - transports anions
Describe F-class ATPases?
AKA ATP synthases
F-class ATPases synthesise ATP using a proton-motive force
Found in the inner membrane of mitochondria, thylakoid membrane of chloroplast and plasma membrane of bacteria
In vivo, F-class ATPases are not powered by ATP - they make it
Describe V-class ATPases?
They transport H+ ions using ATP hydrolysis V-class ATPases are used to lower the pH in intracellular organelles (vacuoles), but are also present in the plasma membrane of some animals (e.g. insect) gut cells They are responsible for bringing H+ ions into the lysosome - as a very acidic pH is required
They hydrolyse ATP in the hydrophilic domain, a ‘rotary’ movement in the hydrophobic domain results in proton transport - across the membrane, against their concentration gradient
What do vacuoles contain other than V-class ATPases?
Vacuoles also contain chloride channels (ClC) to prevent the build up of electrochemical potentials Chloride channels (CLC) might be proton-chloride - transported together to lower pH
Describe P-class ATPases?
They undergo phosphorylation as they transport cations across a membrane
Types:
Proton pump, Na2+/K+ pump, H+/K+, Ca2+ pump, metal pump and flippases
They can be used to detoxify metals like Cu+, Ag+, Zn2+, Cd2+or Pb2+
It is not always confirmed as dimer
There is also a lipid transporter P-type ATPase
Describe the structure of the P-class pumps?
Contains one or two identical a-subunits containing:
Transmembrane ‘pump’ - where the pumping mechanism occurs
Phosphorylation site
ATPase domain - where ATP hydrolysis occurs
Actuator domain - regulatory function for the protein
There is sometimes a Beta domain - which can also be used as a regulatory domain
Describe the P-class Ca2+ ATPase?
Calcium is very important for homeostasis of Ca2+ during muscle contraction
There are two states E1 and E2 that are regulated by phosphorylation
Under physiological conditions almost all intermediates have nucleotide attached - either as ADP or as ATP
What is the mechanism of the P-class Ca2+ ATPase?
- Calcium and ATP binding - Ca2+-ATP bind to the E1 conformational state
- Phosphorylation of aspartate - the ATP is hydrolysed and the Pi phosphorylates aspartate on the E1 conformation of the pump
Here with have an occluded state, the channel from the inside/outside for Ca2+ is essentially closed - Conformational change - E1 changes to E2P (as it is still phosphorylated) and ATP binds
- Calcium release - Calcium is released and exchanged/protonated with 1-3 H+ = H-E2P-ATP
Number of H+ depends on the conditions of the cell - Dephosphorylation - the phosphate on aspartate is then released = H-E2-ATP
- Conformational change - the pump relaxes back to E1, where the Ca2+ and ATP binding sites aren’t occluded
Describe the conformational sites E1 and E2 in the P-class Ca2+ ATPase?
E1 the side is open to the cytosol where Ca2+ can bind down the channel
E2 is open to the (lumen) or other compartment
E1 and E2 states are not defined to what side the protein is open but are defined on their affinities for Ca and H
E1 - high affinity for Ca2+ = 2 bound Ca2+
E2 - low affinity for Ca2+ = no bound Ca2+
Phosphorylation drives the conversion of E1 -> E2 states, providing energy for the uphill transport of Ca2+
Describe the P-class Na2+/K+ ATPase?
It is a heterodimer of catalytic (alpha) and glycoprotein (beta) subunits
It moves 3 Na+ out and K+ in (Antiport)
Coupled with ATP hydrolysis
It has two conformational states E1 and E2
E1 - the binding site has a higher affinity for Na+ and lower affinity for K+
E2 - the binding site has a lower affinity for Na+ and a higher affinity for K+
Located in the plasma membrane of all animal cells
It is a member of a family of ion-translocating ATPases that share highly homologous catalytic subunits
What is the mechanism of P-class Na2+/K+ ATPase (steps 1-3)?
- Na+ and ATP binding - Na+-ATP bind to the E1 conformation = 3Na-E1-ATP
- Phosphorylation of aspartate - the ATP is hydrolysed and the Pi phosphorylates aspartate on the E1 conformation of the pump = 3Na-E1P-ADP
Here with have an occluded state, the channel from the inside/outside for Ca2+ is essentially closed - Conformational change - there is a change in the state from E1 to E2 and due to this change the binding site for K+ has become available = 3Na-E2P
What is the mechanism of P-class Na2+/K+ ATPase (steps 4-7)?
- Na2+ release and K+ binding - Here 3Na+ is released and 2K+ bind, as well as ATP binding = 2K-E2P-ATP
- Dephosphorylation and conformational change - Pi is released which induces E2 back to E1 = 2K-E1P-ATP - with another occluded state
- K+ release - K+ is released on the opposite side to Na2+ as it is in the E1 state
- Na+ and ATP are free to bind again
The binding site of K+ and 2 of the Na+ overlap (same AA involved)
What are ABC transporters?
There are 7 types
They pump - ions, sugars, amino acids and other polar and non-polar substances
Built from 4 molecules:
2 highly conserved cytoplasmic nucleotide-binding domains - binding ATP
2 transmembrane domains - where transport occurs
They are responsible for drug resistance
Further describe the structure of ABC transporters?
4 molecules
2 - nucleotide binding domains
2 - transmembrane domains
Based on TMD sequence and architecture, 7 distinct folds or ‘types’ can be identified
Four domains are typically expressed as: Four peptides (tetramer, bacterial import) Two peptides (dimer, bacterial export or in mitochondria) One peptide (mammalian export)
