Expression, Purification, and Characterization of IP3-binding Domain from Human Type 2

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Nakhaei Amroudie M.; ATAEI F.

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Journal: JOURNAL OF BIOTECHNOLOGY Year:2018 | Volume:9 | Issue:2 Page(s): 165-169

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Information Journal Paper

Title

Expression, Purification, and Characterization of IP3-binding Domain from Human Type 2

Author(s)

Nakhaei Amroudie M. | ATAEI F. | Issue Writer Certificate

Keywords

IP3-binding ReceptorQ1

ExpressionQ1

Intrinsic FluorescenceQ1

LigandQ1

Abstract

Aims: IP3 is a key regulator molecule in the message transmission pathway, and releases calcium into the cytoplasm by binding intracellular IP3R receptors on the surface of the internal calcium stores. The aim of this study was Expression, purification, and characterization of IP3-binding domain from human type 2. Materials & Methods: In this experimental study, the pET-28a plasmid of the carrier of the IP3BD gene was transferred to the E. coli Expression strain BL21 (DE3) by chemical method. In order to optimize the Expression in the bacterial system, the Expression was studied in different conditions, and various temperatures such as 16, 18, 20, and 24° C, the different times after incubation, type of inducer, and its different concentrations were investigated. The induced bacteria were purified on the basis of thermal shock through nickel column for chromatography and the purity of the protein was measured through SDSPAGE. The fluorescence emission of IP3-binding domain was measured in the presence and absence of an IP3 Ligand at wavelength of 295nm. Findings: Protein did not have a significant Expression in LB, TB, and 2xYT environments, and no changes were observed at different times. Expression of bacterial protein at 20° C based on thermal shock of 42° C was higher than in all cases. The purification of the induced bacteria was difficultly repeated due to thermal shock, and the purified samples did not have high concentrations. The fluorescence emission of the protein decreased in the presence of the IP3 Ligand. Conclusion: The bacterial Expression of IP3-binding domain from human type 2 is weak, but the Expression of protein increases with the induction of shock of 42° C.

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APA: Copy

Nakhaei Amroudie, M., & ATAEI, F.. (2018). Expression, Purification, and Characterization of IP3-binding Domain from Human Type 2. JOURNAL OF BIOTECHNOLOGY, 9(2 ), 165-169. SID. https://sid.ir/paper/231281/en

Vancouver: Copy

Nakhaei Amroudie M., ATAEI F.. Expression, Purification, and Characterization of IP3-binding Domain from Human Type 2. JOURNAL OF BIOTECHNOLOGY[Internet]. 2018;9(2 ):165-169. Available from: https://sid.ir/paper/231281/en

IEEE: Copy

M. Nakhaei Amroudie, and F. ATAEI, “Expression, Purification, and Characterization of IP3-binding Domain from Human Type 2,” JOURNAL OF BIOTECHNOLOGY, vol. 9, no. 2 , pp. 165–169, 2018, [Online]. Available: https://sid.ir/paper/231281/en

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