BB15 Membrane Protein Pumps

Question 0

2 types of ATP-driven pumps

Answer 0

• P-type ATPases

* ATP-binding cassette (ABC) transporters

Question 1

Ion pumps are

Answer 1

• energy transducers – convert 1 form of free energy into another

Question 2

ABC transporters undergo

Answer 2

conformational changes on ATP binding and hydrolysis and cause a bound ion to be transported across the membrane

Question 3

Ion pumps are

Answer 3

gradient driven

Question 4

Secondary transporter

Answer 4

• 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

Question 5

Expression and Metabolic Activity

Answer 5

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

Question 6

A transport process must be

Answer 6

• active when deltaG is (+)

* passive when deltaG is (-)

Question 7

Free energy stored in concentration gradient

for uncharged solute molecule

Answer 7

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

Question 8

Free energy stored in concentration gradient

for charged solute molecule

Answer 8

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)

Question 9

Concentration of K+ and Na+ in animal cells

Answer 9

• high K+
• low Na+
relative to external medium

Question 10

Na+ and K+ gradients generated by

Answer 10

Na+ - K+ ATPase
• 3 Na+ out
• 2 K+ in
for each ATP hydrolyzed

Question 11

… provides the energy needed to pump Na+ out of the cell and K+ into the cell, generating gradients

Answer 11

ATP hydrolysis

Question 12

Calcium pump structure

Answer 12

• SR Ca2+ ATPase = SERCE
• P-type ATPase
• forms phosphorylaspartate

Question 13

E1 – Ca2+ bound state

Answer 13

• pumps Ca into SR of muscle cells = important for muscle contraction
• N – binds nucleotide
• P (Asp-351) – accepts phosphoryl group
• A – actuator domain

Question 14

E2 – Ca2+ free state

Answer 14

• calcium binding sites disrupted
• N & P domains closed around phosphorylaspartate analog
• calcium access from cytoplasmic site

Question 15

Mechanism of P-type ATPases

Answer 15

1. (Ca2+)2 binding from cytoplasm
   • E1 to E1-(Ca2+)2
2. ATP binding
   • E1-(Ca2+)2 (ATP)
3. ATP cleavage with transfer of a phosphoryl group to ASP-351 on the enzyme
   E1 – P - (Ca2+)2 – (ADP)
4. ADP release, eversion of enzyme to release CA2+ on the opposite side of the membrane (inside)
   E2-P
5. Hydrolysis of the phosphorylaspartate residue
   • E2
6. Eversion to prepare for the binding of Ca2+ from cytoplasm
   • E1
   •• binding of 2 Ca ions from cyto side of membrane completes cycle

Question 16

…inhibits the Na+ - K+ pump

Answer 16

digitalis (foxglove)

• by blocking dephosphorylation of E2-P

Question 17

Digitoxigenin

Answer 17

• used to treat congestive heart failure

* increases the force of muscle contraction

Question 18

How inhibition of the sodium-potassium pump leads to stronger contraction of the heart

Answer 18

• 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

Question 19

ABC transporters

Answer 19

• 2 transmembrane domains
• 2 ATP-binding domains (cassettes)
• multi-drug resistance (MDR) – pumps drug out

Question 20

ATP transporter mechanism

Answer 20

1. opening of the channel toward the inside of the cell
2. substrate binding and conformational change in the ATP-binding cassettes
3. ATP binding and further conformational changes
4. separation of the membrane-binding domains and release of the substrate to the other side of the membrane (outside)
5. ATP hydrolysis to reset the transporter to its original state
   (opens to inside, drug in, ATP in, conformational change, drug to outside, ATP hydrolysis)

Question 21

Lactose permease

Answer 21

• 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

Question 22

Permease

Answer 22

1. binds a proton from outside of the cell (COO- to COOH)
2. binds substrate from outside
3. eversion
4. releases proton inside
5. releases substrate inside
6. everts to complete the cycle

Question 23

Vibrio cholerae lipid transporter

Answer 23

ABC transporter