Aptamer-based downstream processing of his-tagged proteins utilizing magnetic beads.

Aptamers are synthetic nucleic acid-based high affinity ligands that are able to capture their corresponding target via molecular recognition. Here, aptamer-based affinity purification for His-tagged proteins was developed. Two different aptamers directed against the His-tag were immobilized on magnetic beads covalently. The resulting aptamer-modified magnetic beads were characterized and successfully applied for purification of different His-tagged proteins from complex E. coli cell lysates. Purification effects comparable to conventional immobilized metal affinity chromatography were achieved in one single purification step. Moreover, we have investigated the possibility to regenerate and reuse the aptamer-modified magnetic beads and have shown their long-term stability over a period of 6 months.

Kökpinar Ö ，Walter JG ，Shoham Y ，Stahl F ，Scheper T ... -  《-》

A new application of aptamer: One-step purification and immobilization of enzyme from cell lysates for biocatalysis.

Aptamers are nucleic acid-based high affinity ligands that are able to capture their corresponding target through molecular recognition. In this study, several DNA aptamers with high affinity and specificity for β-glucuronidases (PGUS-E) were obtained by our modified SELEX method. Among them, Apt5 and Apt9 were selected as representatives and covalently linked to magnetic beads, respectively. The aptamer-modified magnetic beads were characterized and successfully applied to one-step purification and immobilization of PGUS-E from the complex cell lysates. By conveniently adjusting the pH and ion strength, the PGUS-E purities reached 84% for Apt5-modified beads and 88% for Apt9-modified beads. Moreover, the maximum PGUS-E capturing capacity of the Apt5 and Apt9 modified magnetic beads were found to be 31.75μg/mg and 32.95μg/mg, respectively. The immobilized PGUS-E on aptamer-based magnetic beads showed good reusability, and the conversion of glycyrrhizin still remained more than 70% after 7 cycles. In addition, the aptamer-modified beads support can be easily regenerated, and the conversion rate of glycyrrhizin (GL) was still 62% after the 7th cycle of regeneration. This investigation can be easily extended to other enzyme systems and may help open a generic route to develop a novel enzyme immobilization technology for biocatalysis based on aptamer.

Qiao L ，Lv B ，Feng X ，Li C ... -  《-》

Systematic investigation of optimal aptamer immobilization for protein-microarray applications.

Walter JG ，Kökpinar O ，Friehs K ，Stahl F ，Scheper T ... -  《-》

Aptamer-modified magnetic beads in affinity separation of proteins.

Zhu G ，Walter JG 《-》

Single-step purification of recombinant Thermus aquaticus DNA polymerase using DNA-aptamer immobilized novel affinity magnetic beads.

Oktem HA ，Bayramoglu G ，Ozalp VC ，Arica MY ... -  《-》