Human ceruloplasmin. Intramolecular electron transfer kinetics and equilibration

Area of Science:

• Biochemistry
• Biophysical Chemistry
• Protein Electron Transfer

Background:

• Ceruloplasmin is a multi-copper oxidase involved in iron metabolism and cellular antioxidant defense.
• Understanding its electron transfer (ET) mechanisms is crucial for elucidating its biological functions.
• Intramolecular ET pathways are initiated by reduction of disulfide bridges, forming RSSR(-) radicals.

Purpose of the Study:

• To map the intramolecular electron transfer cascade in ceruloplasmin following disulfide reduction.
• To identify the specific copper centers and disulfide groups involved in these ET processes.
• To determine the kinetics and energetics of these electron transfer events.

Main Methods:

• Pulse radiolysis to induce RSSR(-) radicals.
• Utilizing the known three-dimensional structure of ceruloplasmin.
• Kinetic analysis of electron transfer between disulfide radicals and Type 1 (T1) copper centers.

Main Results:

• The T1A copper center in domain 6 is the initial site of reduction by RSSR(-) from domain 5 (k = 28 s⁻¹).
• Electron equilibration occurs between T1A and the trinuclear copper center (k = 2.9 s⁻¹).
• A subsequent ET process involves RSSR(-) and the T1B copper center in domain 4 (k = 3.9 s⁻¹).

Conclusions:

• The study elucidates a sequential intramolecular electron transfer pathway in ceruloplasmin.
• Domain 6 (T1A) and domain 4 (T1B) copper centers are sequentially reduced.
• The T1C center in domain 2 appears to be in a reduced state in the resting enzyme and may not participate in this ET cascade.