99765 20231027130639.0 doi 10.1155/2021/6673661 sideral 123304 ART-2021-123304 eng Romero-Tamayo, S. W196 and the ß -Hairpin Motif Modulate the Redox Switch of Conformation and the Biomolecular Interaction Network of the Apoptosis-Inducing Factor 2021 Access copy available to the general public Unrestricted The human apoptosis-inducing factor (hAIF) is a moonlight flavoprotein involved in mitochondrial respiratory complex assembly and caspase-independent programmed cell death. These functions might be modulated by its redox-linked structural transition that enables hAIF to act as a NAD(H/+) redox sensor. Upon reduction with NADH, hAIF undergoes a conformational reorganization in two specific insertions - the flexible regulatory C-loop and the 190-202 ß-harpin - promoting protein dimerization and the stabilization of a long-life charge transfer complex (CTC) that modulates its monomer-dimer equilibrium and its protein interaction network in healthy mitochondria. In this regard, here, we investigated the precise function of the ß-hairpin in the AIF conformation landscape related to its redox mechanism, by analyzing the role played by W196, a key residue in the interaction of this motif with the regulatory C-loop. Mutations at W196 decrease the compactness and stability of the oxidized hAIF, indicating that the ß-hairpin and C-loop coupling contribute to protein stability. Kinetic studies complemented with computational simulations reveal that W196 and the ß-hairpin conformation modulate the low efficiency of hAIF as NADH oxidoreductase, contributing to configure its active site in a noncompetent geometry for hydride transfer and to stabilize the CTC state by enhancing the affinity for NAD+. Finally, the ß-hairpin motif contributes to define the conformation of AIF's interaction surfaces with its physiological partners. These findings improve our understanding on the molecular basis of hAIF''s cellular activities, a crucial aspect for clarifying its associated pathological mechanisms and developing new molecular therapies. info:eu-repo/grantAgreement/ES/DGA/ARAID info:eu-repo/grantAgreement/ES/DGA-FEDER/E35-20R info:eu-repo/grantAgreement/ES/MICINN-AEI/PID2019-103901GB-I00 info:eu-repo/grantAgreement/ES/MINECO-AEI-FEDER/BIO2016-75183-P info:eu-repo/semantics/openAccess by http://creativecommons.org/licenses/by/3.0/es/ 7.31 2021 CELL BIOLOGY 55 / 195 = 0.282 2021 Q2 T1 1.233 2021 Biochemistry 2021 Q1 Medicine (miscellaneous) 2021 Q1 Cell Biology 2021 Q1 9.3 2021 info:eu-repo/semantics/article info:eu-repo/semantics/publishedVersion Laplaza, R. Velazquez-Campoy, A. Universidad de Zaragoza (orcid)0000-0001-5702-4538 Villanueva, R. (orcid)0000-0001-6676-9478 Medina, M. Universidad de Zaragoza (orcid)0000-0001-8743-0182 Ferreira, P. Universidad de Zaragoza (orcid)0000-0003-4076-6118 1002 060 Universidad de Zaragoza Dpto. Bioq.Biolog.Mol. Celular Área Bioquímica y Biolog.Mole. 2021 (2021), 6673661 [19 pp] Oxidative Medicine and Cellular Longevity Oxidative Medicine and Cellular Longevity 1942-0900 1775979 http://zaguan.unizar.es/record/99765/files/texto_completo.pdf Versión publicada 2344364 http://zaguan.unizar.es/record/99765/files/texto_completo.jpg?subformat=icon icon Versión publicada oai:zaguan.unizar.es:99765 articulos driver 2023-10-27-12:53:36 ARTICLE