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- ItemCharacterization of Fluorescent Proteins with Intramolecular Photostabilization*(Weinheim : Wiley-VCH, 2021) Henrikus, Sarah S.; Tassis, Konstantinos; Zhang, Lei; van der Velde, Jasper H. M.; Gebhardt, Christian; Herrmann, Andreas; Jung, Gregor; Cordes, ThorbenGenetically encodable fluorescent proteins have revolutionized biological imaging in vivo and in vitro. Despite their importance, their photophysical properties, i. e., brightness, count-rate and photostability, are relatively poor compared to synthetic organic fluorophores or quantum dots. Intramolecular photostabilizers were recently rediscovered as an effective approach to improve photophysical properties of organic fluorophores. Here, direct conjugation of triplet-state quenchers or redox-active substances creates high local concentrations of photostabilizer around the fluorophore. In this paper, we screen for effects of covalently linked photostabilizers on fluorescent proteins. We produced a double cysteine mutant (A206C/L221C) of α-GFP for attachment of photostabilizer-maleimides on the β-barrel near the chromophore. Whereas labelling with photostabilizers such as trolox, a nitrophenyl group, and cyclooctatetraene, which are often used for organic fluorophores, had no effect on α-GFP-photostability, a substantial increase of photostability was found upon conjugation to azobenzene. Although the mechanism of the photostabilizing effects remains to be elucidated, we speculate that the higher triplet-energy of azobenzene might be crucial for triplet-quenching of fluorophores in the blue spectral range. Our study paves the way for the development of fluorescent proteins with photostabilizers in the protein barrel by methods such as unnatural amino acid incorporation. © 2021 The Authors. ChemBioChem published by Wiley-VCH GmbH
- ItemSynthesis, Characterization, and Physicochemical Performance of Nonionic Surfactants via PEG Modification of Epoxides of Alkyl Oleate Esters(Washington, DC : ACS Publ., 2023) Ogunjobi, Joseph K.; Farmer, Thomas J.; Clark, James H.; McElroy, Con RobertThe synthesis of surfactants from fatty acid esters via epoxide chemistry has been known for its accompanying challenges, which usually involve the use of toxic homogeneous catalysts in the ring-opening reaction step and generation of many side reaction products. This paper presents environmentally benign routes to a library of nonionic surfactants via a three-step synthesis involving transesterification of methyl oleate to alkyl oleates, epoxidation of the oleate alkene, and solventless heterogeneously catalyzed ring opening of the epoxides with poly(ethylene glycols) of varying chain length under a short reaction time (60 min). The processes were highly atom efficient and afforded a minimum surfactant yield of 80% with limited or negligible side reaction products. The intermediate molecules and synthesized surfactants were purified and comprehensively characterized, including physicochemical measurements: dynamic surface tension and equilibrium surface tension. Additionally, the hydrophilic-lipophilic balance (HLB) concept was used to comprehensively scan through the polarity behaviors of the surfactants’ head and tail in solution as a prediction of their end use. The results showed that surfactants have a critical micelle concentration (CMC) lower than 0.1 mg/ml as the alkyl oleate increases in length from ethyl to decyl and that the lower-molecular-weight surfactants reached equilibrium faster than the higher-molecular-weight surfactants. HLB results showed that the surfactants can be applied as oil-in-water emulsifiers, detergents, solubilizers, and wetting agents. In general, the synthesized surfactants potentially possess switchable properties for use in industrial formulations, as the alkyl chain length and the ethylene oxide number in the surfactant’s structure are varied.