Large-scale production and structural and biophysical characterizations of the human hepatitis B virus polymerase

Journal of Virology
Judit VörösNeil Ferguson

Abstract

Hepatitis B virus (HBV) is a major human pathogen that causes serious liver disease and 600,000 deaths annually. Approved therapies for treating chronic HBV infections usually target the multifunctional viral polymerase (hPOL). Unfortunately, these therapies--broad-spectrum antivirals--are not general cures, have side effects, and cause viral resistance. While hPOL remains an attractive therapeutic target, it is notoriously difficult to express and purify in a soluble form at yields appropriate for structural studies. Thus, no empirical structural data exist for hPOL, and this impedes medicinal chemistry and rational lead discovery efforts targeting HBV. Here, we present an efficient strategy to overexpress recombinant hPOL domains in Escherichia coli, purifying them at high yield and solving their known aggregation tendencies. This allowed us to perform the first structural and biophysical characterizations of hPOL domains. Apo-hPOL domains adopt mainly α-helical structures with small amounts of β-sheet structures. Our recombinant material exhibited metal-dependent, reverse transcriptase activity in vitro, with metal binding modulating the hPOL structure. Calcomine orange 2RS, a small molecule that inhibits duck HBV POL activi...Continue Reading

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Citations

Nov 3, 2014·Annual Review of Virology·Todd M GrecoIleana M Cristea
Sep 10, 2014·Proceedings of the National Academy of Sciences of the United States of America·Christian KönigerMichael Nassal
May 13, 2014·Current Opinion in Virology·Luis Menéndez-AriasBeatriz Pacheco
Oct 16, 2016·Clinics in Liver Disease·Sebastien BoucleRaymond F Schinazi
Aug 1, 2019·Current Drug Targets·Bing-Yi ZhangXiao-Ping Yu
Sep 25, 2019·Antiviral Research·Enzo TramontanoLuis Menéndez-Arias

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