resumo
RNA is a biopolymer of high relevance in the biopharmaceuticals field and in fundamental and applied research; however, preservation of RNA stability is still a remarkable challenge. Herein, we demonstrate the enhanced potential of aqueous solutions of self-buffering cholinium-based Good's buffers ionic liquids (GB-ILs), at 20 and 50% (w/w), as alternative preservation media of recombinant small RNAs. The thermal stability of RNA is highly enhanced by GB-ILs, with an increase of 14 degrees C in the biopolymer melting temperature-the highest increase observed to date with ILs. Most GB-ILs investigated improve the stability of RNA at least up to 30 days, both at 25 degrees C and at 4 degrees C, without requiring the typical samples freezing. Molecular dynamics simulations were applied to better understand the molecular-level mechanisms responsible for the observed RNA improved stability. The number of IL cations surrounding the RNA chain is similar, yet with differences found for the IL anions, which are responsible for the overall charge of the biopolymer first solvation sphere. No cytotoxicity of the studied solutions containing RNA and ILs at 20% (w/w) was observed onto two distinct human cell lines, reinforcing their potential to act as preservation media when foreseeing biopharmaceutical applications. Finally, RNA was successfully recovered from the ILs aqueous solutions, without changes in its structural integrity, and the ILs were successfully recycled and reused.
palavras-chave
DNA; PURIFICATION; SYSTEMS; GREEN
categoria
Chemistry; Science & Technology - Other Topics; Engineering
autores
Pedro, AQ; Pereira, P; Quental, MJ; Carvalho, AP; Santos, SM; Queiroz, JA; Sousa, F; Freire, MG
nossos autores
Augusto Pedro
Investigador Júnior
Mara Guadalupe Freire Martins
Investigador Coordenador
Marcos André Pinto de Carvalho
Bolseiro de investigação
Maria João Quental
Bolseiro de pós-Doutoramento
Sérgio Miguel Fernandes dos Santos
Investigador AuxiliarGrupos
G1 - Materiais Porosos e Nanossistemas
G5 - Materiais Biomiméticos, Biológicos e Vivos
G6 - Materiais Virtuais e Inteligência Artificial
agradecimentos
We acknowledge the CICS-UBI Projects Pest-OE/SAU/UI0709/2014 and UID/Multi/00709/2013, the program COMPETE, Pest-C/SAU/UI0709/2011, and the CICECO-Aveiro Institute of Materials projects, POCI-01-0145-FEDER-007679 (UID/CTM/50011/2013), financed by national funds through the FCT/MEC, and when appropriate, co-financed by FEDER under the PT2020 Partnership Agreement. CICS-UBI was also supported by FEDER funds through the POCI-COMPETE 2020-Operational Programme Competitiveness and Internationalisation in Axis I-Strengthening research, technological development, and innovation (Project POCI-010145-FEDER-007491). We acknowledge the financial support from the European Union Framework Programme for Research and Innovation HORIZON 2020, under TEAMING Grant Agreement No 739572 The Discoveries CTR P.P. acknowledges a postdoctoral fellowship from a FCT-funded project (Reference PTDC/1399/2014), and M.J.Q the FCT Ph.D. Grant SFRH/BD/100155/2014. M.G.F. acknowledges the European Research Council (ERC) for the Starting Grant ERC-2013-StG-337753.