Detection of human annexin A1 as the novel N-terminal tag for separation and purification handle

• Published in: Microbial cell factories 2023-01, Vol.22 (1), p.2-2, Article 2
• Main Authors: He, Xiaomei, Zhang, Shuncheng, Dang, Dongya, Lin, Tingting, Ge, Yuanyuan, Chen, Xiaofeng, Fan, Jun
• Format: Article
• Language: English
• Subjects: Affinity purification; Affinity separation; Annexin A1; Annexin A1 - metabolism; Annexins; Chemical properties; Chromatography, Affinity - methods; Detection; Edetic Acid - metabolism; Escherichia coli; Escherichia coli - metabolism; Fusion tag; Green Fluorescent Proteins - metabolism; Humans; Proteins; Recombinant Fusion Proteins - genetics; Recombinant Fusion Proteins - metabolism; Separation
• ISSN: 1475-2859; 1475-2859
• DOI: 10.1186/s12934-022-02005-x

Several fusion tags for separation handle have been developed, but the fused tag for simply and cheaply separating the target protein is still lacking. Separation conditions for the human annexin A1 (hanA1) tagged emerald green fluorescent protein (EmGFP) in Escherichia coli were optimized via precipitation with calcium chloride (CaCl ) and resolubilization with ethylenediamine tetraacetic acid disodium salt (EDTA-Na ). The HanA1-EmGFP absorbing with other three affinity matrix was detected, only it was strongly bound to heparin Sepharose. The separation efficiency of the HanA1-EmGFP was comparable with purification efficiency of the His6-tagged HanA1-EmGFP via metal ion affinity chromatography. Three fluorescent proteins for the EmGFP, mCherry red fluorescent protein and flavin-binding cyan-green fluorescent protein LOV from Chlamydomonas reinhardtii were used for naked-eye detection of the separation and purification processes, and two colored proteins including a red protein for a Vitreoscilla hemoglobin (Vhb), and a brown protein for maize sirohydrochlorin ferrochelatase (mSF) were used for visualizing the separation process. The added EDTA-Na disrupted the Fe-S cluster in the mSF, but it showed little impact on heme in Vhb. The selected five colored proteins were efficient for detecting the applicability of the highly selective hanA1 for fusion separation and purification handle. The fused hanA1 tag will be potentially used for simple and cheap affinity separation of the target proteins in industry and diagnosis.