Regulation of cellular pH Flashcards
What is the pH of the plasma?
7.4
What is the pH inside the cell?
7-7.1
What scale is pH? (equation)
What does this mean?
Logarithmic scale:
pH = -log[H+]
1 small change in H+ has a LARGE change in pH
How do proteins buffer pH?
1) H+ binds to protein - changes the protein charge
2) Conformational change in the protein
3) Change in protein function
Why can increase in H+ concentration potentially have disastrous effects?
H+ binds to proteins and changes their charge and therefore shape and FUNCTION
What 2 methods can be used to measure intracellular pH?
1) Microelectrodes
2) Fluorescent indicators
Which method is good for measuring pH of large cells?
Describe this method
Microelectrodes:
1) 2 voltage electrodes - measure ion current across the membrane
2) Measure voltage difference between V1 and V2
3) Change in voltage is proportional to pH
How are the electrodes in the microelectrode technique calibrated?
1) Buffer at pH 6 and measure voltage
2) Buffer at pH 8 and measure voltage
3) Construct a curve
How can voltage be converted into pH?
Using the curve constructed:
pH = (V-offset) / slope
Which method is good for measuring pH of small cells?
Describe this method
Fluorescent indicators:
1) Cells loaded with LIPID SOLUBLE, INACTIVE form of the indicator
2) Inside the cell - the inactive form is converted into an active form
3) Indicator is excited with light of a specific wavelength and the amount of EMITTED fluorescence light is measured (emits at a different wavelength)
4) Fluorescence is proportional to intracellular pH and [H+]
In the fluorescent indicator method to measure pH, why is the cell loaded with an INACTIVE form of the indicator?
It is positively charged - so it can cross the membrane
In the fluorescent indicator method to measure pH, how is the inactive form of the indicator turned into the active form?
What does this cause?
Inactive form is cleaved
Active form has a negative charge - is TRAPPED inside the cell
How is the fluorescent indicator method calibrated?
At the end of the experiment:
- Proton ionophore which calibrates pH inside of the cell with the pH outside
- When change pH outside of the cell, pH inside the cell becomes the same
- Look at the signal intensity when the pH changes
What 3 things are involved in the control of intracellular pH?
1) Buffering
2) Acid extrusion
3) Acid loading
What is the relationship between acid extrusion and acid loading in a cell under normal conditions?
What do these systems do?
They are balanced
These systems act to REVERSE pH changes
