Human mitochondrial AAA+ ATPase SKD3/CLPB assembles into nucleotide-stabilized dodecamers.

Area of Science:

• Biochemistry
• Molecular Biology
• Structural Biology

Background:

• SKD3 (human CLPB) is an AAA+ ATPase implicated in 3-methylglutaconic aciduria type VII, congenital neutropenia, and acute myeloid leukemia drug resistance.
• Its precise biological function and mechanism of action, particularly its role in the mitochondrial intermembrane space, remain largely uncharacterized.

Purpose of the Study:

• To elucidate the oligomeric state and structural organization of SKD3 in relation to its nucleotide-binding activity.
• To investigate the structural basis for SKD3's function and its link to associated clinical phenotypes.

Main Methods:

• Sedimentation equilibrium and dynamic light scattering were employed to determine SKD3's oligomeric state.
• Negative-stain electron microscopy (EM) with image-class analysis was used to visualize the structure of nucleotide-bound SKD3.

Main Results:

• SKD3 exists as a monomer at low concentrations but forms oligomers at higher concentrations or upon binding adenine nucleotides.
• Nucleotide-bound SKD3 self-associates into a dodecamer, visualized by EM as cylinder-shaped particles with a central channel.
• The dodecamer appears to be composed of two hexameric rings, a 'double-hexamer sandwich' structure, which is uncommon among AAA+ ATPases.

Conclusions:

• SKD3 forms a unique double-hexameric structure dependent on nucleotide binding.
• This non-canonical structure may be crucial for SKD3's biological functions and its involvement in disease pathogenesis.
• Further research is needed to determine the functional significance of this unusual quaternary structure.