OVERPRODUCTION, PURIFICATION, AND CHARACTERIZATION X PROTEIN HEPATITIS B VIRUS FUSED THIOREDOXIN IN ESCHERICHIA COLI ROSETTA-GAMI
Hepatitis B virus (HBV) cause 96% of hepatitis mortality worldwide. HBV infection leads to chronic hepatitis progression into cirrhosis and liver carcinoma. HBV X protein (HBx) is known to contribute to the pathogenesis of liver carcinoma. HBx structure consist 4 disulfide bonds and 53.7% hydroph...
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| Format: | Theses |
| Language: | Indonesia |
| Online Access: | https://digilib.itb.ac.id/gdl/view/48682 |
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| Institution: | Institut Teknologi Bandung |
| Language: | Indonesia |
| Summary: | Hepatitis B virus (HBV) cause 96% of hepatitis mortality worldwide. HBV
infection leads to chronic hepatitis progression into cirrhosis and liver carcinoma.
HBV X protein (HBx) is known to contribute to the pathogenesis of liver
carcinoma. HBx structure consist 4 disulfide bonds and 53.7% hydrophobic amino
acids causes HBx expression in Escherichia coli as inclusion bodies (IB). The aim
of this study to overproduce, solubilize, renaturate, and purify HBx fused
Thioredoxin (Trx-HBx). In this study, Trx-HBx was expressed in E. coli Rosettagami at low temperatures. Overproduction of protein was optimized in various
induction temperature (17°C and 25°C) and various induction time (8 and 16
hours). The protein was characterized by SDS-PAGE, dot blot, and Western blot.
The results of this study showed that the majority of Trx-HBx was expressed as IB
under optimum temperature at 25°C and 16 hours induction. Trx-HBx was
solubilized from IB using 6 M urea solubilization buffer. Solubilized Trx-HBx was
renatured simultaneous with purification using affinity chromatography nickel
column. Trx-HBx was purified using 200 mM imidazole at the fourth and fifth
elution, but SDS-PAGE reducing and non-reducing did not show disulfide bond
formation after renaturation.
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