High‐efficiency Ni²⁺‐NTA/PAA magnetic beads with specific separation on His‐tagged protein

Abstract

To effective capture and universal enrichment of His-tagged protein, polyacrylic acid (PAA) brushes were used to encapsulate Fe 3 O 4 nanoparticles, connect NTA, and Ni 2+ to prepare magnetic beads. These materials provide many advantages, such as excellent stability, tuneable particle size, and a surface for further functionalisation with biomolecules. Histagged green fluorescence protein (GFP) was separated efficiently, and the binding capacity of Fe 3 O 4 /MPS@PAA/NTA-Ni 2+ was 93.4 mg/g. Compared with High-Affinity Ni-NTA Resin and Ni-NTA Magnetic Agarose Beads, Fe 3 O 4 /MPS@PAA/NTA-Ni 2+ nanocomposites exhibited higher separation efficiency and binding capacity towards His-tagged GFP. Moreover, the selectivity and recyclability of them for the target proteins were maintained well after six cycles. This study would widen the application of PAA in constructing multifunctional nanocomposites for biomedical fields.

Alginate-Based Applications in Biotechnology with a Special Mention to Biosensors

“…His-tag is a sequential hexahistidine residue that can chelate metal ions such as Ni (II), Co (II), Zn (II), and Cu (II). These metal ions can be prepared for immobilisation by treatment with a chelating moiety such as nitrilotriacetic acid [102,103] or iminodiacetic acid and can be used alone. For example, because alginate is polyanionic, several alginate nickel composites have been prepared from alginate and NiCl 2 [104,105].…”

Magnetic Dendritic Polymer Nanospheres for High-Performance Separation of Histidine-Rich Proteins

“…For example, Wang and his co-workers prepared highefficiency Ni 2+ -NTA/PAA magnetic beads with specific separation on His-tagged protein and the binding capacity was 93.4 mg/g. 6 Bai and co-workers reported the preparation of core−shell structured Fe 3 O 4 @SiO 2 superparamagnetic microspheres as immobilized metal ion affinity adsorbents for His-protein purification, and the maximum adsorption capacity for lysozyme was 51.0 mg/g. 36 However, the disadvantages of low surface metal-ion density and poor recyclability have greatly limited these strategies.…”

“…Magnetic nanospheres have been widely used in environmental, biomedical, , and clinical fields , owing to their many unique properties and desirable features, such as ease of synthesis and operation, large surface area, fast separation, good biocompatibility, and recyclable performance, making them the most promising materials in protein isolation and purification. , There are a variety of methods that have been used to prepare Fe 3 O 4 nanoparticles, such as thermal decomposition, , co-precipitation, , microemulsion , and hydrothermal method, , etc., for many applications. Recently, different magnetic-based materials have been prepared and applied to protein purification.…”

“…To develop multifunctional magnetic materials, much work has been done recently. For example, Wang and his co-workers prepared high-efficiency Ni 2+ -NTA/PAA magnetic beads with specific separation on His-tagged protein and the binding capacity was 93.4 mg/g . Bai and co-workers reported the preparation of core–shell structured Fe 3 O 4 @SiO 2 superparamagnetic microspheres as immobilized metal ion affinity adsorbents for His-protein purification, and the maximum adsorption capacity for lysozyme was 51.0 mg/g .…”

Triagem biológica de inibidores de sirtuína 2 (>i<Tc>/i<Sir2rp1) candidatos a antichagásicos

“…Dessa maneira, a proteína marcada com His foi purificada por resina de alta afinidade, contendo esferas de agarose magnéticas de Ni-NTA. Assim, as esferas ligadas à proteína do estudo foram lavadas cinco vezes com tampão de lise contendo 20 mM 40 mM, 60 mM e 300 mM de imidazol, no qual permitiu a separação das proteínas contaminantes da TcSir2rp1 (GUTIÉRREZ et al, 2018;WANG et al, 2020). Além disso, devido a utilização da cepa com chaperonas, foi necessário adicionar mais uma etapa de purificação, a cromatografia de troca iônica em coluna MonoQ ® 5/50 com gradiente de 0 -1 M de NaCl.…”