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Logo 22nd ISBC & 20th ISLS


Application of chemiluminescence and bioluminescence to nanomaterials

Plasmon-induced Increased Absolute Activity of Amydetes and chimeric ZZ-Amy Firefly Luciferases bound to PEG(SH)2- and Cys-Functionalized Gold Nanoparticles

Iseli L. Nantes1, Gabriel F. Pelentir2, Adrianne M. M. Brito3, Alexandre J. Lanfredi4, Vadim Viviani5, Elisângela B.1

1UNIVERSIDADE FEDERAL DO ABC, Centro de Ciências Naturais e Humanas, Brazil, 2Universidade Federal de São Carlos, Depto de Física, Química e Matemática, Campus Sorocaba, Brazil, 3UNIVERSIDADE FEDERAL DO ABC, Centro de Ciências Naturais e Humanas, Brazil., 4UNIVERSIDADE FEDERAL DO ABC, Centro de Ciências Exatas e Sociais Aplicadas, 5Universidade Federal de São Carlos, Campus Sorocaba, Depto de Física, Química e Matemática, Brazil

E-mail: ilnantes@gmail.com

The novel bioluminescent fusion protein, which was formed by the green-blue emitting Amydetes vivianii firefly luciferase (Amy-WT) linked with the N-terminal ZZ portion of protein A (chimeric ZZ-Amy), has been efficiently applied in immun blots for detection of SARS-CoV-2 nucleoprotein. The chimeric ZZ-Amy displays a slightly higher and more sustained luminescent signal when compared to commercial cheiluminescent HRP-labeled secondary antibodies. The increase of Pyrearinus termitilluminans (PyLuc) and Phrixotrix red emitting (RELuc) luciferases stability when associated with gold nanoparticles (AuNPs) capped with dithiol polyethylene glycol (PEG(SH)2AuNPs) inspired similar Nano/Bio construction for Amy-WT and chimeric ZZ-Amy luciferases. The association of the enzymes with the PEG(SH)2AuNPs did not affect the enzymatic structure, as attested by FTIR analysis and bioluminescence spectra. Scanning electron microscopy images showed 10 nm AuNPs aggregated in a dense PEG(SH)2 corona that became rarefied in the presence of Amy-WT. Contrary to the observed for PyLuc and RELuc, Amy-WT and chimeric ZZ-Amy associated with PEG(SH)2AuNPs exhibited a significant luminescence increase (absolute activity) with a slight stability increase. The spectral overlap of luciferases with the absorption band of the 10 nm AuNPs maximizes the effect of confined plasmons on the optical density of states near the emission peaks of Amy-WT and ZZ-Amy. The Nano/ZZ-Amy construction improves the detection limits of the bioluminescence for several practical applications.

Keywords: luciferases, gold nanoparticles, plasmon resonance, ZZ-Amy.

Acknowledgments: Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP): 2017/02317-2, 2022/04800-0; CAPES 001; Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq) 2021/313111-9 , 405060/2022-1

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