Defining NADH-Driven Allostery Regulating Apoptosis-Inducing Factor.

Area of Science:

• Biochemistry
• Cell Biology
• Structural Biology

Background:

• Apoptosis-inducing factor (AIF) plays a vital role in mitochondrial respiratory complex assembly and necroptotic cell death.
• AIF's functions are regulated by allosteric switching, influenced by NADH charge-transfer complex (CTC) formation.
• The precise molecular mechanisms underlying AIF allostery remain incompletely understood.

Purpose of the Study:

• To elucidate the molecular pathways connecting AIF's active site to its allosteric switching regions.
• To characterize how NADH CTC formation triggers allosteric changes in AIF.
• To provide a structural and dynamic basis for AIF's flavoswitch mechanism.

Main Methods:

• Characterization of dimer-permissive AIF mutants using small-angle X-ray scattering (SAXS) and X-ray crystallography.
• Molecular dynamics (MD) simulations to probe communication networks between AIF and the CTC.
• Biochemical assays to validate identified allosteric pathways.

Main Results:

• Two distinct pathways mediating allostery from the CTC active site to AIF were identified.
• Pathway 1 involves active site residue H454 linking to S480 in AIF's central beta-strand, modulating the dimerization interface.
• Pathway 2 involves an interaction network including the FAD cofactor, central beta-strand, and C-loop, where R529 reorientation facilitates C-loop release during CTC formation.

Conclusions:

• The study defines specific molecular pathways responsible for AIF allosteric regulation.
• Understanding these pathways, particularly the flavoswitch mechanism, is key to AIF's role in mitochondrial homeostasis and cell death.
• This knowledge offers potential targets for biomedical interventions aimed at controlling AIF activity.