Design of a miniaturized FeS4 protein

Despite the great success of computational protein design in recent years,[1,2] the structure/function correlation of iron-binding proteins designed from scratch remained elusive. Explicitly, the rational arrangement of the first and second coordination interactions able to purposely select and stabilize the very labile iron chemistry is still a difficult task[3]. Besides in repurposed natural scaffolds or by exogenous ligands like porphyrins and polypyridines[4], iron-bound de novo proteins have never been structurally characterized before. We present, for the first time, the structural and functional features of a fully designed FeS4 protein, namely METPsc. Inspired by natural rubredoxins, this miniaturized protein does not hold any sequence correlation to the known congeners, as assessed by BLASTP. Strikingly, METPsc 28-long sequence stores all the information required to fold around the metal in a tetrahedral geometry and to function as an electron-transfer protein, as confirmed by crystallography, UV-Vis and EPR spectroscopy, and cyclic voltammetry. Finally, we exploited its terminal electron acceptor properties in an artificial electron chain triggered by visible light. Its applicability in optoelectronics and light-harvesting biodevices is being explored. [1] F. Pirro, N. Schmidt, J. Lincoff, Z. X. Widel, N. F. Polizzi, L. Liu, M. J. Therien, M. Grabe, M. Chino, A. Lombardi, W. F. DeGrado, Proc. Natl. Acad. Sci. 2020, 117, 33246–33253. [2] Z. Chen, R. D. Kibler, A. Hunt, F. Busch, J. Pearl, M. Jia, Z. L. Van Aernum, B. I. M. Wicky, G. Dods, H. Liao, M. S. Wilken, C. Ciarlo, S. Green, H. El-Samad, J. Stamatoyannopoulos, V. H. Wysocki, M. C. Jewett, S. E. Boyken, D. Baker, Science 2020, 368, 78–84. [3] E. N. Mirts, A. Bhagi-Damodaran, Y. Lu, Acc. Chem. Res. 2019, 52, 935–944. [4] F. Nastri, D. D’Alonzo, L. Leone, G. Zambrano, V. Pavone, A. Lombardi, Trends Biochem. Sci. 2019, 44, 1022–1040.