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The N-Terminal 6×His Tag on β-Clamp Processivity Factor Occludes Gly66 and Affects the Growth of Escherichia coli B834 (DE3) Cells

  • MOLECULAR CELL BIOLOGY
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Abstract

The affinity tags in fusion proteins are extensively used in protein expression techniques. The most common affinity tags, such as glutathione S-transferase (GST), poly-histidine, maltose binding protein (MBP), and streptavidin tags, are routinely used for increasing expression, improving solubility, and facilitating protein purification. The large affinity tags (MBP, GST) are known to influence the conformational homogeneity and, therefore, the three-dimensional structure of in vivo folded proteins. The current study described in vivo effects of small affinity fusion 6×His tag on the growth of cells. Hexa-histidine tagged full length β-clamp and non-hexa-histidine tagged β-clamp were over-expressed and co-expressed in possible combinations with truncated DnaE in E. coli expression strain. After the induction with IPTG, the protein expression was assessed by SDS PAGE. The comparative analysis of the growth curves generated for the induced and un-induced cells demonstrated a decrease in growth rates of the cells over-expressing non-6×His tagged β-clamp as compared to 6×His tagged β-clamp. Based on the analysis of the soluble and insoluble protein fractions by SDS PAGE gels and published His-tagged β-clamp structure (PDB: 4K74) we propose that N-terminal 6×His Tag on β-clamp occludes its Gly66 to ultimately affect its ability to interact with the δ subunit of the clamp loader.

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Authors and Affiliations

  1. Institute of Microbiology, University of Sindh, 71000, Jamshoro, Pakistan

    A. A. Patoli & B. B. Patoli

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  1. A. A. Patoli
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  2. B. B. Patoli
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Correspondence to A. A. Patoli or B. B. Patoli.

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The text was submitted by the author(s) in English.

Place of Study: School of Biology, Queens Medical Centre, University of Nottingham, Nottingham, NG7 2UH, UK.

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Patoli, A.A., Patoli, B.B. The N-Terminal 6×His Tag on β-Clamp Processivity Factor Occludes Gly66 and Affects the Growth of Escherichia coli B834 (DE3) Cells. Mol Biol 53, 32–37 (2019). https://doi.org/10.1134/S0026893319010126

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