1 research outputs found
Key Role of the Adenylate Moiety and Integrity of the Adenylate-Binding Site for the NAD<sup>+</sup>/H Binding to Mitochondrial Apoptosis-Inducing Factor
Apoptosis-inducing factor (AIF) is
a mitochondrial flavoprotein
with pro-life and pro-death activities, which plays critical roles
in mitochondrial energy metabolism and caspase-independent apoptosis.
Defects in AIF structure or expression can cause mitochondrial abnormalities
leading to mitochondrial defects and neurodegeneration. The mechanism
of AIF-induced apoptosis was extensively investigated, whereas the
mitochondrial function of AIF is poorly understood. A unique feature
of AIF is the ability to form a tight, air-stable charge-transfer
(CT) complex upon reaction with NADH and to undergo a conformational
switch leading to dimerization, proposed to be important for its vital
and lethal functions. Although some aspects of interaction of AIF
with NAD<sup>+</sup>/H have been analyzed, its precise mechanism is
not fully understood. We investigated how the oxidized and photoreduced
wild-type and G307A and -E variants of murine AIF associate with NAD<sup>+</sup>/H and nicotinamide mononucleotide (NMN<sup>+</sup>/H) to
determine the role of the adenylate moiety in the binding process.
Our results indicate that (i) the adenylate moiety of NAD<sup>+</sup>/H is crucial for the association with AIF and for the subsequent
structural reorganization of the complex, but not for protein dimerization,
(ii) FAD reduction rather than binding of NAD<sup>+</sup>/H to AIF
initiates conformational rearrangement, and (iii) alteration of the
adenylate-binding site by the G307E (equivalent to a pathological
G308E mutation in human AIF) or G307A replacements decrease the affinity
and association rate of NAD<sup>+</sup>/H, which, in turn, perturbs
CT complex formation and protein dimerization but has no influence
on the conformational switch in the regulatory peptide