CLONING, EXPRESSION AND CHARACTERIZATION OF RECOMBINANT HUMAN FC RECEPTOR LIKE 1, 2 AND 4 MOLECULES

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SHABANI MAHDI; HEMMATI AZAM; ZANDEMAMI MAHDI; KHOSHNOODI JALAL; JEDDI TEHRANI MAHMOOD; RABBANI HODJATALLAH; AMIRGHOFRAN ZAHRA; SHOKRI FAZEL

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Journal: Iranian Journal of Biotechnology Year:2013 | Volume:11 | Issue:3 (43) Page(s): 182-192

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Title

CLONING, EXPRESSION AND CHARACTERIZATION OF RECOMBINANT HUMAN FC RECEPTOR LIKE 1, 2 AND 4 MOLECULES

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Keywords

FCRLQ2

HIS-TAGQ2

POLYCLONAL ANTIBODYQ2

PROTEIN EXPRESSIONQ3

PURIFICATIONQ2

Abstract

Background: The Fc receptor like (FCRL) molecules belong to the immunoglobulin (Ig) superfamily with potentially immunoregulatory function. Among the FCRL family FCRL2 and 4 are predominantly expressed on memory B cells and FCRL1 is a pan- B cell marker. To date, no ligand has been identified for the human FCRL1, 2 and 4 molecules.Objectives: Cloning, expression, PURIFICATION and structural analysis of the extracellular domain of human FCRL1, 2 and 4 proteins.Materials and Methods: In this study, the extracellular part of human FCRL1, 2 and 4 were subcloned into prokaryotic expression vectors pET-28b (+) and transformed into BL21-DE3 E.coli strain. PROTEIN EXPRESSION was optimized by fine adjustments such as induction time, incubation temperature and expression hosts. Recombinant FCRL proteins were purified by metal affinity chromatography using Ni-NTA resin. Purified FCRL proteins were further characterized by SDS-PAGE and immunoblotting using HIS-TAG and FCRL specific polyclonal antibodies.Results: Our results demonstrated that FCRL1, 2 and 4 were successfully expressed in pET-28b (+) vector. Optimization of the expression procedure showed that IPTG induction at OD600 = 0.9 and overnight incubation at 37oC resulted in the highest expression levels of FCRL proteins ranging from approximately 15% (FCRL1) to 25% (FCRL2 and 4) of the total bacterial lysate proteins.Conclusions: These purified recombinant proteins are potentially a valuable tool for investigating the immunoregulatory function of FCRL molecules and the production of specific mAbs for immunotherapeutic interventions.

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APA: Copy

SHABANI, MAHDI, HEMMATI, AZAM, ZANDEMAMI, MAHDI, KHOSHNOODI, JALAL, JEDDI TEHRANI, MAHMOOD, RABBANI, HODJATALLAH, AMIRGHOFRAN, ZAHRA, & SHOKRI, FAZEL. (2013). CLONING, EXPRESSION AND CHARACTERIZATION OF RECOMBINANT HUMAN FC RECEPTOR LIKE 1, 2 AND 4 MOLECULES. IRANIAN JOURNAL OF BIOTECHNOLOGY, 11(3 (43)), 182-192. SID. https://sid.ir/paper/139259/en

Vancouver: Copy

SHABANI MAHDI, HEMMATI AZAM, ZANDEMAMI MAHDI, KHOSHNOODI JALAL, JEDDI TEHRANI MAHMOOD, RABBANI HODJATALLAH, AMIRGHOFRAN ZAHRA, SHOKRI FAZEL. CLONING, EXPRESSION AND CHARACTERIZATION OF RECOMBINANT HUMAN FC RECEPTOR LIKE 1, 2 AND 4 MOLECULES. IRANIAN JOURNAL OF BIOTECHNOLOGY[Internet]. 2013;11(3 (43)):182-192. Available from: https://sid.ir/paper/139259/en

IEEE: Copy

MAHDI SHABANI, AZAM HEMMATI, MAHDI ZANDEMAMI, JALAL KHOSHNOODI, MAHMOOD JEDDI TEHRANI, HODJATALLAH RABBANI, ZAHRA AMIRGHOFRAN, and FAZEL SHOKRI, “CLONING, EXPRESSION AND CHARACTERIZATION OF RECOMBINANT HUMAN FC RECEPTOR LIKE 1, 2 AND 4 MOLECULES,” IRANIAN JOURNAL OF BIOTECHNOLOGY, vol. 11, no. 3 (43), pp. 182–192, 2013, [Online]. Available: https://sid.ir/paper/139259/en

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