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Expression and purification of soluble bacterially-expressed human hexokinase II in E.coli system

Tanbin, Suriyea and Ahmad Fuad, Fazia Adyani (2019) Expression and purification of soluble bacterially-expressed human hexokinase II in E.coli system. In: "9th International Conference on Biomedical Engineering and Technology, ICBET 2019", 28 - 30 Mar 2019, Meiji UniversityTokyo, Japan.

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Human hexokinase II (HKII) is one of the key enzymes in the glycolytic pathway. It has been postulated that HKII is a potential target for anti-dengue (DENV) drug development, as well as involved in cancer and tumor cell growth. In this work, the human hexokinase II (HKII) gene was cloned into pETite N-His SUMO vector and transformed into the E.coli strain HI-control 10G for the propagation of clones. Two different expression hosts, E.coli HI-controlTM BL21 (DE3) and BL21 (DE3) pLysS were used to optimize HKII expression. In order to obtain the soluble recombinant HKII in a functional form, we optimized protein expression at three different temperatures; 17°C, 25°C and 37°C, at 24 hours incubation time. The soluble protein was expressed in the presence of 0.5 mM isopropyl-2-Dthiogalactopyranoside (IPTG) in TB media at 17°C for 24 hrs. The expressed protein was then purified to homogeneity by a combination of Immobilized Metal Ion Affinity Chromatography (IMAC), size exclusion chromatography (SEC) and ion-exchange chromatography (IEX), resulting in pure bacterially-expressed HK2. Taken together, this study has successfully produced soluble bacterially-expressed human HKII that can be utilized for further therapeutic studies.

Item Type: Conference or Workshop Item (Slide Presentation)
Additional Information: 7200/79728
Uncontrolled Keywords: Glycolysis; Human hexokinase IIProtein expressionProtein purification
Subjects: Q Science > QH Natural history
T Technology > TP Chemical technology > TP248.13 Biotechnology
Kulliyyahs/Centres/Divisions/Institutes (Can select more than one option. Press CONTROL button): Kulliyyah of Engineering > Department of Biotechnology Engineering
Depositing User: Dr Fazia Adyani Ahmad Fuad
Date Deposited: 24 Jun 2020 12:20
Last Modified: 17 Mar 2022 16:08
URI: http://irep.iium.edu.my/id/eprint/79728

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