Key Role of the Adenylate Moiety and Integrity of the Adenylate-Binding Site for the NAD+/H Binding to Mitochondrial Apoptosis-Inducing Factor
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- 作者：Luca Sorrentino ; Alessandra Maria Calogero ; Vittorio Pandini ; Maria Antonietta Vanoni ; Irina F. Sevrioukova ; Alessandro Aliverti
- 刊名：Biochemistry
- 出版年：2015
- 出版时间：December 1, 2015
- 年：2015
- 卷：54
- 期：47
- 页码：6996-7009
- 全文大小：696K
- ISSN：1520-4995
文摘
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+/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+/H and nicotinamide mononucleotide (NMN+/H) to determine the role of the adenylate moiety in the binding process. Our results indicate that (i) the adenylate moiety of NAD+/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+/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+/H, which, in turn, perturbs CT complex formation and protein dimerization but has no influence on the conformational switch in the regulatory peptide.