BB15 Membrane Protein Pumps Flashcards
2 types of ATP-driven pumps
- P-type ATPases
* ATP-binding cassette (ABC) transporters
Ion pumps are
• energy transducers – convert 1 form of free energy into another
ABC transporters undergo
conformational changes on ATP binding and hydrolysis and cause a bound ion to be transported across the membrane
Ion pumps are
gradient driven
Secondary transporter
- uses the gradient of one ion to drive the active transport of another
- different mechanism of active transport
- eg E. coli lactose transporter
- present in the membranes of our cells
- expression of these transporters determines which cell metabolites a cell can import from the environment –> expression = primary means of controlling metabolism
Expression and Metabolic Activity
eg. glucose metabolism
• which tissues can use glucose is governed by the expression of different members of a family of glucose transporters: GLUT1 – GLUT5 in different cell types
• GLUT3 binds glucose tightly so these cells have first call on glucose when it is present at low concentrations
A transport process must be
- active when deltaG is (+)
* passive when deltaG is (-)
Free energy stored in concentration gradient
for uncharged solute molecule
deltaG = RT ln (c2/c1) deltaG = 2.303 log10 (c2/c1)
- R = gas constant (kJ/mol)
- T = temp in K
- c1 = concent on side 1
- c2 = concent on side 2
Free energy stored in concentration gradient
for charged solute molecule
deltaG = RT ln (c2/c1) + Z F deltaV deltaG = 2.303 RT log10 (c2/c2) + Z F deltaV
- Z = charge of solute
- F = faraday constant (96.5 kJ/V/mol)
Concentration of K+ and Na+ in animal cells
• high K+
• low Na+
relative to external medium
Na+ and K+ gradients generated by
Na+ - K+ ATPase
• 3 Na+ out
• 2 K+ in
for each ATP hydrolyzed
… provides the energy needed to pump Na+ out of the cell and K+ into the cell, generating gradients
ATP hydrolysis
Calcium pump structure
- SR Ca2+ ATPase = SERCE
- P-type ATPase
- forms phosphorylaspartate
E1 – Ca2+ bound state
- pumps Ca into SR of muscle cells = important for muscle contraction
- N – binds nucleotide
- P (Asp-351) – accepts phosphoryl group
- A – actuator domain
E2 – Ca2+ free state
- calcium binding sites disrupted
- N & P domains closed around phosphorylaspartate analog
- calcium access from cytoplasmic site
Mechanism of P-type ATPases
- (Ca2+)2 binding from cytoplasm
• E1 to E1-(Ca2+)2 - ATP binding
• E1-(Ca2+)2 (ATP) - ATP cleavage with transfer of a phosphoryl group to ASP-351 on the enzyme
E1 – P - (Ca2+)2 – (ADP) - ADP release, eversion of enzyme to release CA2+ on the opposite side of the membrane (inside)
E2-P - Hydrolysis of the phosphorylaspartate residue
• E2 - Eversion to prepare for the binding of Ca2+ from cytoplasm
• E1
•• binding of 2 Ca ions from cyto side of membrane completes cycle
…inhibits the Na+ - K+ pump
digitalis (foxglove)
• by blocking dephosphorylation of E2-P
Digitoxigenin
- used to treat congestive heart failure
* increases the force of muscle contraction
How inhibition of the sodium-potassium pump leads to stronger contraction of the heart
- inhibition of the Na+ - K+ pump by digitalis leads to higher level of Na+ inside the cell
- reduced Na+ gradient results in slower extrusion of calcium by the sodium-calcium exchanger
- increase in calcium enhances the ability of the cardiac muscle to contract
ABC transporters
- 2 transmembrane domains
- 2 ATP-binding domains (cassettes)
- multi-drug resistance (MDR) – pumps drug out
ATP transporter mechanism
- opening of the channel toward the inside of the cell
- substrate binding and conformational change in the ATP-binding cassettes
- ATP binding and further conformational changes
- separation of the membrane-binding domains and release of the substrate to the other side of the membrane (outside)
- ATP hydrolysis to reset the transporter to its original state
(opens to inside, drug in, ATP in, conformational change, drug to outside, ATP hydrolysis)
Lactose permease
• gradient – not ATP
• H+ outside = higher concentration, drives uptake of lactose/sugars against concentration gradient
• archetype secondary transporter
• the thermodynamically unfavourable flow of one species of ion/molecule UP a concentration gradient is driven by the favourable flow of a different species DOWN a concentration gradient
- antiporter = diff molecules, diff directions
- symporter = diff molecules, same directions
- uniporter = same molecule, diff directions
Permease
- binds a proton from outside of the cell (COO- to COOH)
- binds substrate from outside
- eversion
- releases proton inside
- releases substrate inside
- everts to complete the cycle
Vibrio cholerae lipid transporter
ABC transporter
