Competition Between Protein and DNA for Binding to Natural Sepiolite Nanofibers

David Adame Brooks,1,2 Olivier Piétrement,3 Elodie Dardillac,1 Fidel Antonio Castro Smirnov,4 Pilar Aranda,5 Eduardo Ruiz-Hitzky,5 Bernard S Lopez1 1Université de Paris Cité, INSERM U1016, UMR 8104 CNRS, Institut Cochin, Paris, France; 2Centro de Biofísica Médica (Cbiomed), Universidad de Oriente, Santiago de Cuba, Cuba; 3Université Bourgogne Europe, CNRS, Laboratoire interdiciplinaire Carnot de Bourgogne ICB UMR 630, Dijon, F-21000, France; 4Instituto Superior de Tecnologías Ciencias Aplicadas, Universidad de la Habana (Instec-UH), Quinta de Los Molinos, Ave. Salvador Allende Luaces, Habana, 6163, Cuba; 5Instituto de Ciencia de Materiales de Madrid (ICMM), CSIC, c/Sor Juana Inés de la Cruz 3, Madrid, 28049, SpainCorrespondence: Bernard S Lopez, Institut Cochin 24 rue du Faubourg St Jacques, Paris, 75014, France, Email [email protected]: Sepiolite nanofibers, which are natural silicates belonging to the clay mineral family, could be promising potential nanocarriers for the nonviral transfer of biomolecules. The physicochemical characteristics of sepiolite make it capable of binding various types of biological molecules, including polysaccharides, lipids, proteins and viruses. Sepiolite nanofibers have also been shown to bind effectively to various types of DNA molecules through electrostatic interactions, hydrogen bonds, cationic bridges and van der Waals forces. In this study, we analyzed the adsorption of DNA and proteins to sepiolite by analyzing the competition among these biomolecules during the adsorption process.Methods: To determine the binding of sepiolite to proteins, we used BSA and a monoclonal antibody (mAb) against the CD4 membrane antigen as a model. The binding efficiency was measured by adsorption isotherms. Zeta potential measurements of the suspensions were performed using a Brookhaven NanoBrook 90 Plus PALS instrument.Results: We show here that the adsorption of proteins to sepiolite is increased in the presence of CaCl2 and is charge-dependent and that sepiolite can adsorb proteins even when their net charges are equal to those on its surface. Coating of sepiolite with DNA (Sep/DNA bionanocomposites) reduces the absorption efficiency of both BSA and mAb, and this can be rescued by CaCl2. Conversely, preincubation of sepiolite with BSA or the mAb decreased the efficiency of DNA binding; Ca2+ restored the binding efficiency for BSA but not for the mAb. Changes in pH result in changes in the net charge of proteins, influencing the amount of protein adsorbed.Conclusion: Although various types of protein interactions with mineral clays have been described, our results confirm that electrostatic forces are among the primary interactions in the adsorption process. These results pave the way for the use of biohybrids as a new class of nanoplatform for gene transfer with potential clinical applications.Keywords: bionanomaterials, sepiolite clay mineral, sepiolite-DNA interaction, sepiolite-protein interaction