Evaluation of induction conditions for multi-antigen chimeric TgAGS/BsT protein from Toxoplasma gondii in Escherichia coli BL 21 Star during batch cultivation.

Abstract

The present study evaluated the induction conditions (inducer type and concentration) for the expression of the multi-antigenic chimeric protein TgAGS/BsT from Toxoplasma gondii in Escherichia coli BL21 Star during batch submerged cultivation. With a molecular mass of about 18.6 kDa, the chimeric TgAGS/BsT protein has potential use in a diagnostic kit and as a vaccine candidate for toxoplasmosis. Batch cultures were performed at 37°C and induced by the addition of different concentrations of isopropyl β-D-1-thiogalactopyranoside (IPTG) (0.1, 1.0, and 2.0 mM) and lactose (1.0, 10.0, and 20.0 g/L) in different culture media (2xTY and Terrific Broth). The parameters used to analyze the cultures were cell growth, acetic acid formation, pH variation, plasmid stability, and protein expression evaluated by SDS-PAGE, together with spectrophotometric techniques. Higher concentrations of IPTG (2.0 mM) and lactose (10.0 and 20.0 g/L) promoted higher acetic acid accumulation and a continuous decrease in pH, indicating metabolic stress. The pH profile varied with the medium: 2xTY showed partial recovery due to acetate reabsorption, while TB exhibited a continuous decline. Plasmid stability remained high (>90%) across all evaluated conditions. The results demonstrate that the concentration and type of inducer, combined with medium composition, have a strong influence on the balance between cell growth, metabolism, and the efficiency of heterologous protein expression. Lactose induction provided a more gradual process and achieved higher volumetric protein concentrations in TB medium (0.039 g/L with 10.0 g/L lactose) and higher protein yield per biomass in 2xTY medium (0.0086 g/g with 10.0 g/L lactose), whereas increasing IPTG concentrations did not enhance protein accumulation.