Question

1. What is the significance of peak area around 500nm in cytochrome c spectrum? Under what conditions two splits peaks vs. one round peak in this region are observed?

2. Which component of cytochrome c is responsible for the red color and a clear absorption spectrum of cytochrome c?

3. If the ration of I/I₀ is equal to 0.01, calculate the %T and A values for this.

4. A sample of pure cytochrome c was diluted by taking 100microL and adding 900microL of buffer. Calculate the concentration of cytochrome c inmM, and in mg/mlof original sample with an absorption value of 0.236 in a cell of 1 cm path length using equation 2 in the lab protocol and table for molar absorptivities. What is the total cytochrome c in mg if the total volume fraction is 7.5mL? The mol.wt of cytochrome c is 12,300 g/mol.

Answer 1

cytochrome c oxidase is involved in the electron transport system and is the terminal member of the respiratory chain. It consists of two heme (heme 'a' and 'a₃' ) and two Cu (Cu A and Cu B) units.

Heme a and Cu 'B' are involved in the proton pumping process whereas heme a₃ and Cu 'A' are involved in oxygen binding.

At around 500nm the peak area in the spectrum indicates the binding of oxygen atom with the metal centre in the cytochrome c oxidase biomolecule. Splitting of peak into two is because of the difference in the oxidation state of the two iron atoms in the heme group as both iron are low spin but with (+2) and (+3) oxidation states.

Red colour and a clear absorption spectrum of cytochrome c oxidase is due to the intraligand charge transfer of electrons from pie bonding to pie antibonding of the porphyrin ring.