Lecture 5

Question 1

pH equation

Answer 1

pH = -log10[H+]

Question 2

Why does pH need to be so carefully controlled?

Answer 2

Proteins act to buffer changes in H+
Change in protein charge
Change in protein conformation
Change in protein function
DISASTER

Question 3

H+ increase

Answer 3

Compensation = remove H+/ alkalinisation

Question 4

H+ decrease

Answer 4

Compensation = increase H+/acidify

Question 5

Measuring intracellular pH with micro electrodes

Answer 5

Using 2 micro electrodes V1 and V2 containing a H+ sensitive resin
Difference between two electrodes measured
Change in voltage proportional to change in pH

Question 6

V1

Answer 6

Measures all components of the normal current

Question 7

V2

Answer 7

Measures potential for all ions except protons

Question 8

voltage vs. pH graph

Answer 8

V=slopexpH

pH=(v-offset)/slope

Question 9

Measuring intracellular pH using fluorescent indicators

Answer 9

Cells loaded with lipid soluble inactive form of indicator
Inside cell the indicator is converted into the active form(-ve)
Indicator is excited with light with a specific wavelength
the amount of fluorescence at a second wavelength is measured
Fluorescence is proportional to intracellular pH
Indicator calibrated inside of cell
Membrane is permeabilised with a proton ionophore and the pH of the bath solution is charged
In presence of the ionophore the bath pH is the same as the intracellular pH

Question 10

Factors involved in pH control

Answer 10

Buggering
Acid extrusion
Acid loading

Question 11

pH buffer

Answer 11

Any system that moderates the effects of an acid or an alkali load by reversibly consuming or releasing protons

Question 12

Buffering

Answer 12

Systems act to minimise pH changes and help protect the cell from damage
Do not prevent pH changes, just minimise magnitude of the change
Can’t reverse pH changes

Question 13

Buffer power

Answer 13

Defined as the amount of strong base that must be added to a solution in order to raise the pH by a given amount

Question 14

protein buffering

Answer 14

Relies on:
COOH is a proton donor
NH2 is a proton receiver
Found on amino acid residues

Question 15

COOH group

Answer 15

pH increases = loss of proton = = pH decrease

Question 16

NH2 group

Answer 16

pH decreases = gain of proton = increase in pH

Question 17

Acid Extrusion

Answer 17

uses NHE proton
Na+ into cell (relies on Na+/K+ ATPase)
H+ out of the cell –> Raising the pH

Question 18

NHE setpoint

Answer 18

When more alkaline than set point = NHE inactive

Activation at an acidic pH

Question 19

NHE allosteric modification

Answer 19

Protons other than the ones being transported can bind to the protein causing a conformational change which increases its activity
Acidic pH = higher chance of this so increased activity

Question 20

NHE-1

Answer 20

Has a housekeeping function - regulated intracellular pH and cell volume
Inhibited by low conc of amiloride
Found in basolateral membrane of epithelial cells
2 TM domains

Question 21

Acid loading

Answer 21

Uses Cl-/HCO3- exchanger
Cl- into and HCO3- out of the cell
Removal of HCO3- leaves H+ behind to decrease pH inside cell

Question 22

Cl-/HCO3- activity

Answer 22

Sensitive to pH
Low activity at high pH
High activity at a high pH

Question 23

Anion Exchanger Family

Answer 23

4 subtypes
All isoforms are inhibited by the stilbene derivate drug (DIDS)
Exchange of Cl- and HCO3- independent of Na+
13 TM domains

Question 24

AE1

Answer 24

Predominantly in the red blood cells and kidneys

Responsible for th eChloride/hamburger shift

Question 25

Maintaining a steady state pH

Answer 25

When JL=JE
No net proton flux
Gives the resting intracellular pH

Question 26

JE

Answer 26

Rate of acid extrusion

Question 27

JL

Answer 27

Rate of acid loading