Defining NADH-Driven Allostery Regulating Apoptosis-Inducing Factor

Abstract

SUMMARY Apoptosis-inducing factor (AIF) is critical for mitochondrial respiratory complex biogenesis and for mediating necroptotic parthanatos: these functions are seemingly regulated by enigmatic allosteric switching driven by NADH charge-transfer complex (CTC) formation. Here we define molecular pathways linking AIF’s active site to allosteric switching regions by characterizing dimer-permissive mutants using small-angle X-ray scattering (SAXS) and crystallography and by probing AIF-CTC communication networks using molecular dynamics simulations. Collective results identify two pathways propagating allostery from the CTC active site: 1) active site H454 links to S480 of AIF’s central β-strand to modulate a hydrophobic border at the dimerization interface and 2) an interaction network links AIF’s FAD cofactor, central β-strand, and Cβ-clasp whereby R529 reorientation initiates C-loop release during CTC formation. This knowledge of AIF allostery and its flavoswitch mechanism provides a foundation for biologically understanding and biomedically controlling its participation in mitochondrial homeostasis and cell death.

The MRE11–RAD50–NBS1 Complex Conducts the Orchestration of Damage Signaling and Outcomes to Stress in DNA Replication and Repair

“…Yet, this requires a path for a comprehensive characterization of proteins and nucleic acids, including their biologically relevant complexes and conformations (223). These observations have motivated our efforts to develop high-throughput and objective X-ray methods for comprehensive measurements of complexes and conformations and flexibility under near-physiological conditions (e.g., see 224–229). Thus, X-ray scattering and diffraction methods coupled to cryo-EM and single-molecule data are helping to define the MRN structural biochemistry described here.…”

Small angle X‐ray scattering and cross‐linking for data assisted protein structure prediction in CASP 12 with prospects for improved accuracy

“…SAXS provides multiple measures of flexibility that merit consideration for inclusion in computational prediction: lack of convergence in Kratky plot, invalidation Porod‐Debye law, Flory's inequality, low particle density, and inability to model data with a single model. Combining protein volume derived from SAXS data with measured mass allows calculation of the SAXS‐based density, which can vary significantly for flexible proteins: flexible proteins are likely to have densities of 0.9‐1.0 g·cm −3 , which is far below the canonical value of 1.37 g·cm −3 for folded compact proteins . In fact, there is a huge opportunity for computational prediction methods that employ SAXS data and incorporate the bound hydration layer and local flexibility, as these 2 features result in all crystal structures having substantially more error in the refined crystallographic models than in the measured diffraction data .…”

Targeting Allostery with Avatars to Design Inhibitors Assessed by Cell Activity: Dissecting MRE11 Endo- and Exonuclease Activities

“…Unlike other structural techniques, SAXS enables comprehensive assessment of conformation and assembly under near physiological conditions (Hura et al, 2013). So we can expect it to be increasingly used for ligand-binding assessments and their impact on assembly, as seen for the apoptosis-inducing factor that links mitochondrial metabolism, DNA damage responses, and programmed cell death (Brosey et al, 2016). …”