Jessica Cecilia Berrones-Reyes

Designing photochemical properties in proteins is a frontier challenge in biomolecular design, which, when realised, will open new avenues in catalysis and biomolecular electronics. Here we demonstrate an integrated computational design framework, supported by ultrafast spectroscopy, combining protein design, quantum mechanical/molecular mechanics molecular dynamics (QM/MM MD) and electronic structure calculations, to control flavin excited states in de novo proteins. Iterative design yields FC-4D2, a flavocytochrome with covalently bound flavin and non-covalently bound heme B and Fl-4D2, where the heme-binding site is removed. Both proteins are highly thermostable, and access all physiologically relevant flavin redox and protonation states, while high-resolution X-ray crystallography confirms the near-Ångström-level accuracy in design. Incorporating QM/MM MD and linear-response time-dependent density functional theory (LR-TDDFT) into the design workflow enables engineering of a flavin–tryptophan charge-transfer variant, Fl-4D2 _CT , that modifies the flavin absorption spectrum, substantially alters its redox properties and suppresses fluorescence. Ultrafast studies reveal how protein design reshapes the excited state relaxation timescale and mechanism. Our study establishes a modular strategy for programming flavin photochemistry in designed proteins.

Designing redox proteins with predictable and tuneable electron transfer properties is a major goal in de novo bioenergetics. Here we show that replacing heme B with a series of structurally conservative non natural metalloporphyrins enables broad modulation of redox potentials over 400 mV in the de novo designed monoheme m4D2 and diheme 4D2 T19D. The non-natural porphyrins bind with high affinity and do not compromise either the heme binding site or global protein structure, as evidenced by X-ray crystallography and NMR spectroscopy. We also report the native-like NMR structure of m4D2 loaded with the non-natural and symmetric iron 2,4-dimethyldeuteroporphyrin IX, confirming our modular approach to tetrahelical redox protein design. This work establishes a versatile platform for constructing tuneable electron carriers for engineered bioenergetic pathways and bioelectronic applications.

The fluorescence imaging technologies are becoming the most powerful and noninvasive diagnostic tools in cellular biology and modern medicine where abnormal cell arrangements are associated with diseases. Thus, these techniques require new fluorescent dyes with excellent chemical, physical, and photophysical properties. A series of four new Boron Schiff bases (1–4) has been prepared by condensation between phenylboronic acid with the corresponding ligand. The compounds were characterized by NMR (¹H, ¹³C, and ¹¹B), UV/vis, fluorescence spectroscopy, and high‐resolution mass spectrometry. The crystal structures of three compounds showed tetracoordinated Boron atoms with semiplanar skeleton ligands. Interesting organoboron response to viscosity on their fluorescence (Φ: more than 3‐fold). Additionally, compounds 1 and 2 were found to serve as a fluorescent dye for cell imaging (B16F10, CaCo, and A‐431 cells) since it has the capability to rapidly accumulate within the cells and gave bright green fluorescence, it showed low cytotoxicity activity and high photostability in solution. Additionally, the compounds have also been investigated using DFT.

A series of eight new organotin compounds derived from Schiff bases has been prepared by a multicomponent reaction from 2-hydroxy-1-naphthaldehyde or 4-substituted-2-hydroxybenzalhedyde, benzhydrazine, and the corresponding diorganotin oxide (R₂SnO, R = ⁿBu or Ph). All of the compounds were fully characterized by NMR (¹H, ¹³C, and ¹¹⁹Sn), IR, UV/vis, elemental analyses and fluorescence spectroscopy. The crystal structures for some organotin compounds were determined by single crystal X-ray diffraction analysis. All of the compounds display fluorescence at room temperature with quantum yields of about 2 × 10⁻⁴ to 0.56. The cytotoxic activity and cellular imaging studies were carried out with the newly synthesized compounds. To the best of our knowledge, this is the first report of organotin compounds with Schiff base ligands investigated for fluorescence bioimaging (FBI).