Lecture 12 - AQP continued

Question 1

Mercurial agents work by

Answer 1

binding to cysteine residues

Question 2

AQP 1 has 4 cysteine residues at positions…

Answer 2

87, 102, 152 and 189

Question 3

AQP1 cysteine mutated to serine

Answer 3

don’t bind mercury anymore but have the same water permeability as in WT

Question 4

C189S

Answer 4

mercury has no effect on this mutant so can conclude this is where mercury binds

Question 5

Cys189 location

Answer 5

C in loop E of the AQP1 hourglass model

So it is in the pore

Question 6

AQP1 pores

Answer 6

where the loops overlap when subunits join together to form a pore

Question 7

AQP1 structure

Answer 7

tetramer with 4 small water pores and a large big pore in the middle

Question 8

Tandem dimers fore water pore experiments

Answer 8

half WT half MT, water permeability reduced by 50% so proves each monomeric subunit is a water pore

Question 9

AQP1 water transport

Answer 9

channels are water selective and proton-impermeable
As water moves through the channel, hydrogen bonds between H2O molecules are broken so it is like a selectivity filter through the pore
interact with asparagine residue to prevent protons

Question 10

AQP1 and asparagine motifs

Answer 10

H2O binds to asparagine in the pore to prevent protons coming through

Question 11

mercury inhibition experiment

Answer 11

mutated equivalent Cys189s residue in bacterial AQP residue, making in mercury-sensitive
found mercury binds in the middle of each water pore

Question 12

AQP3 location

Answer 12

basolateral membrane of collecting duct and generally throughout the body

Question 13

AQP3 permeability

Answer 13

water, glycerol and urea

Question 14

AQP3 and glycerol

Answer 14

mercury inhibits glycerol permeability which suggests that it goes through the monomeric pore

Question 15

AQP6 location

Answer 15

intracellular

Question 16

AQP6 and mercury

Answer 16

mercury stimulated water permeability so it activated AQP6

Question 17

AQP6 permeability and acidic conditions

Answer 17

acidic conditions stimulate chloride conductance and water permeability

Question 18

AQP6 histone residue

Answer 18

linked to pH sensitivity, when histone is lost there is a loss of pH sensitivity

Question 19

AQP2 and diabetes insipidus

Answer 19

mutation near channel pore results in reduced H2O channel permeability

Question 20

AQP1 and the proximal tubule

Answer 20

high water permeability

AQP1 contributes 70%