Differences in functional cross-talk between loops C and D in two mitochondrial cytochromes

Area of Science:

• Biochemistry
• Molecular Biology
• Protein Science

Background:

• Cytochrome c (cyt c) heme is shielded by variable loop sequences across species.
• Human (hu) cyt c mutations G41S and Y48H are linked to thrombocytopenia.
• Species-specific sequence variations impact protein structure and function.

Purpose of the Study:

• Investigate the effects of G41S and Y48H mutations in horse heart (hh) cyt c.
• Compare these effects to those observed in hu cyt c.
• Elucidate the role of sequence differences in loop dynamics, electron transfer, and peroxidase activity.

Main Methods:

• Molecular Dynamics (MD) simulations to probe loop dynamics.
• Experimental characterization of the alkaline transition.
• Measurement of reduction potential and intrinsic peroxidase activity.

Main Results:

• Mutations in hh cyt c reduced loop C-D contacts and enhanced loop D dynamics.
• Y48H mutation decreased reduction potential; both mutations increased peroxidase activity.
• Ligand switch rates (k_f) increased in hh cyt c mutants but decreased in hu cyt c mutants.

Conclusions:

• Sequence variations between hu and hh cyt c, particularly residue 46, influence loop interactions and dynamics.
• These differences modulate electron transfer and peroxidase activity.
• Species-specific sequence variations are critical for cyt c functional regulation.