Structural basis for adPEO-causing mutations in the mitochondrial TWINKLE helicase

Area of Science:

• Biochemistry
• Molecular Biology
• Genetics

Background:

• TWINKLE is essential for mitochondrial DNA (mtDNA) replication and maintenance in mammals.
• Mutations in TWINKLE's linker region cause autosomal dominant progressive external ophthalmoplegia (adPEO), a neuromuscular disorder linked to mtDNA deletions.
• The molecular mechanisms underlying adPEO due to TWINKLE mutations are not fully understood, but altered oligomerization is suspected.

Purpose of the Study:

• To characterize adPEO-associated TWINKLE variants using single-particle electron microscopy.
• To elucidate the molecular basis of how TWINKLE mutations lead to diminished helicase activity and adPEO.
• To understand the role of TWINKLE oligomerization and conformational changes in disease pathogenesis.

Main Methods:

• Single-particle electron microscopy (cryo-EM) was used to analyze wild-type and mutant TWINKLE proteins.
• Biochemical assays were performed to assess the activities of TWINKLE variants.
• Conformational changes upon nucleoside triphosphate binding were investigated.

Main Results:

• TWINKLE variants exhibit altered oligomeric properties, correlating with diminished enzymatic activities.
• Mutations were categorized based on their impact on linker flexibility, ring closure, or subunit stoichiometry.
• Wild-type TWINKLE undergoes significant conformational changes upon nucleotide binding, a property lost in disease variants.

Conclusions:

• adPEO-causing TWINKLE mutations disrupt protein oligomerization and/or conformational dynamics, leading to impaired mtDNA maintenance.
• Understanding these molecular defects provides insights into the pathogenesis of adPEO and related mitochondrial disorders.
• This research may inform the development of therapeutic strategies for adPEO and other pathologies linked to TWINKLE dysfunction.