Optimization and scale-up production of Zika virus ?NS1 in Escherichia coli: application of Response Surface Methodology.

ABSTRACT: Diagnosing Zika virus (ZIKV) infections has been challenging due to the cross-reactivity of induced antibodies with other flavivirus. The concomitant occurrence of ZIKV and Dengue virus (DENV) in endemic regions requires diagnostic tools with the ability to distinguish these two viral infections. Recent studies demonstrated that immunoassays using the C-terminal fragment of ZIKV NS1 antigen (?NS1) can be used to discriminate ZIKV from DENV infections. In order to be used in serological tests, the expression/solubility of ?NS1 and growth of recombinant E. coli strain were optimized by Response Surface Methodology. Temperature, time and IPTG concentration were evaluated. According to the model, the best condition determined in small scale cultures was 21 °C for 20 h with 0.7 mM of IPTG, which predicted 7.5 g/L of biomass and 962 mg/L of ?NS1. These conditions were validated and used in a 6-L batch in the bioreactor, which produced 6.4 g/L of biomass and 500 mg/L of ?NS1 in 12 h of induction. The serological ELISA test performed with purified ?NS1 showed low cross-reactivity with antibodies from DENV-infected human subjects. Denaturation of ?NS1 decreased the detection of anti-ZIKV antibodies, thus indicating the contribution of conformational epitopes and confirming the importance of properly folded ?NS1 for the specificity of the serological analyses. Obtaining high yields of soluble ?NS1 supports the viability of an effective serologic diagnostic test capable of differentiating ZIKV from other flavivirus infections.

SUBMITTER: Kanno AI 

PROVIDER: S-EPMC6938527 | biostudies-literature | 2019 Dec

REPOSITORIES: biostudies-literature