Coupling proteins to magnetic nanoparticles for tuning their properties by magnetic heating

Resumen del póster presentado al 5th Multistep Enzyme Catalyzed Processes Congress (MECPC), celebrado online del 13 al 16 de septiembre de 2021.It is well known that magnetic nanoparticles (MNPs) absorb magnetic energy when exposed to an alternating magnetic field (AMF) and therefore dissipate heat in their local environment. So far, the heat dissipated by the MNPs has been widely used in biomedicine for cancer treatment and drug delivery. The FET-OPEN project HOTZYMES proposes instead to exploit this property for the biotechnological production of pharmaceuticals and bio-commodities, by conjugating thermophilic enzymes on the MNPs surface in order to selectively tune their activity by triggering this highly localized magnetic heating while maintaining the reaction media at low temperature.To this aim superparamagnetic iron oxide nanoparticles coated with different organic shells (dimercaptusuccinic acid, DMSA; polyacrylic acid, PAA) have been selected due to their good colloidal stability, high density of functionalizable chemical groups (-COOH) and a good combination of different heating efficiencies. They have been further functionalized with NTA-Cu2+, a chelating agent commonly used in protein purification by metal affinity interaction. Herein, we demonstrate that we can selectively bind a His-tag variant of the superfolded GFP protein, thanks to the high affinity of the immobilized copper to the 6xhistidine tag genetically fused to the protein. Furthermore, using the same strategy, we were able to bind the tetrameric recombinant His-tag variant of the B. stearothermophilus alcohol dehydrogenase (ADH) with also high specificity. Additionally, we demonstrate that the application of an AMF to the selected protein@MNPs derivatives can lead to the generation of different temperature gradients on the MNPs surface by studying both the GFP fluorescence (that strongly depends on temperature), and the effect of the triggered local heating on ADH activity. These first results suggest the feasibility of tuning remotely protein properties by combining their coupling to MNPs with the application of alternating magnetic fields.The research for this work has received funding from the European Union (EU) project HOTZYMES (grant agreement n° 829162) under EU’s Horizon 2020 Programme Research and Innovation actions H2020- FETOPEN-2018-2019-2020-01. Authors also thank Spanish MINECO project BIO2017-84246-C2-1-R and DGA and Fondos Feder (Bionanosurf E15_17R).Peer reviewe