Exploring the inhibitor binding pocket of respiratory complex I

Area of Science:

• Biochemistry
• Molecular Biology
• Structural Biology

Background:

• Hydrophobic compounds and detergents inhibit respiratory chain complex I's ubiquinone reductase activity.
• The structure of complex I's peripheral arm from Thermus thermophilus provides a basis for investigating inhibitor binding.

Purpose of the Study:

• To identify residues involved in ubiquinone and inhibitor binding within complex I.
• To elucidate the spatial arrangement of binding sites for different classes of hydrophobic inhibitors.

Main Methods:

• Site-directed mutagenesis in Yarrowia lipolytica targeting the 49-kDa and PSST subunit interface.
• Assessing changes in inhibition (IC50 values) for various hydrophobic inhibitors and detergents.

Main Results:

• Mutations in specific regions altered inhibitor sensitivity, revealing distinct binding sites.
• Overlap was observed between binding sites for DQA (class I) and rotenone (class II) inhibitors.
• A separate region near iron-sulfur cluster N2 was exclusively affected by the detergent C(12)E(8) (type C).

Conclusions:

• Provides structure-based evidence for distinct but overlapping binding sites for hydrophobic inhibitors in complex I.
• Suggests these binding sites may extend to the ubiquinone reduction site.
• Enhances understanding of complex I inhibition mechanisms and structure-function relationships.