DNA binding suppresses human AIF-M2 activity and provides a connection between redox chemistry, reactive oxygen species, and apoptosis.
Human AIF-M2 is an unusual flavoprotein oxidoreductase that binds DNA, nicotinamide coenzyme, and the modified flavin 6-hydroxy-FAD. Using multiple solution methods to investigate the redox chemistry and binding interactions of AIF-M2, we demonstrate that binding of DNA and coenzyme to AIF-M2 is mutually exclusive. We also show that DNA binding does not perturb the redox chemistry of AIF-M2, but it has significant effects on the reduction kinetics of the 6-hydroxy-FAD cofactor by NAD(P)H. Based on quantitative analysis of ligand binding and redox chemistry, we propose a model for the function of AIF-M2. In this model, DNA binding suppresses the redox activity of AIF-M2 by preventing the binding of the reducing coenzyme NAD(P)H. This DNA-mediated suppression of AIF-M2 activity is expected to lower cellular levels of superoxide and peroxide, thereby lessening survival signaling by Ras, NF-kappaB, or AP-1, as suggested from knock-out studies of the related AIF in human colon cancer cell lines. We show marked differences between AIF-M2 and AIF. DNA and coenzyme binding activity is retained in the C-terminal deletion mutant AIF-M2-(Delta319-613), whereas DNA binds to the C-terminal D3 domain of AIF. Our work provides the first analysis of AIF-M2 ligand interactions and redox chemistry and identifies an important mechanistic connection between coenzyme and DNA binding, redox activity, and the apoptotic function of AIF-M2. Through its DNA binding activity, we suggest that AIF-M2 lessens survival cell signaling in the presence of foreign (e.g. bacterial and (retro)viral) cytosolic DNA, thus contributing to the onset of apoptosis.
Gong
Min
M
Manchester Interdisciplinary Biocentre and Faculty of Life Sciences, University of Manchester, M1 7DN Manchester, United Kingdom.
Hay
Sam
S
Marshall
Ker R
KR
Munro
Andrew W
AW
Scrutton
Nigel S
NS
D017209:Apoptosis; D051017:Apoptosis Regulatory Proteins / Q000737:chemistry*; D003600:Cytosol / Q000378:metabolism; D004247:DNA / Q000737:chemistry*; D006801:Humans; D007700:Kinetics; D024101:Mitochondrial Proteins / Q000737:chemistry*; D009154:Mutation; D009243:NAD / Q000737:chemistry; D010084:Oxidation-Reduction; D010088:Oxidoreductases / Q000378:metabolism; D010100:Oxygen / Q000378:metabolism; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D017382:Reactive Oxygen Species / Q000378:metabolism; D015398:Signal Transduction
C462504:ferroptosis suppressor protein 1, human; D051017:Apoptosis Regulatory Proteins; D024101:Mitochondrial Proteins; D017382:Reactive Oxygen Species; D009243:NAD; D004247:DNA; D010088:Oxidoreductases; D010100:Oxygen