Successful design of an industrial biological product such as a vaccine requires an efficient, robust and reproducible process. In this study, we investigated conditions for process optimization of a promising recombinant factor H binding protein (fHbp)–Porin A (PorA) chimeric protein-based vaccine candidate against Meningococcal B serogroup in Escherichia coli B834 strain using the inducible T7-lac promoter system. Random screening of components of complex culture media for growth and expression using IPTG as an inducer, showed inconsistency when analyzed using Plackett–Burman design (PBD). Further analysis identified galactose present either in tryptone (as lactose) or soy-phytone, has caused strong autoinduction and is surmised as an interfering factor for IPTG induction. Synergistic combination of soy-phytone and yeast extract under conditions of controlled growth and auto-induction gave a good correlation between the cell growth and expression of the fHbp–PorA chimeric protein when evaluated using PBD and central composite design. Using response optimization conditions, an optimized media was attained. A shake flask study was performed to validate the optimized media, and later, a fed-batch fermentation at 10 L scale was established to prove the scalability, consistency, and product quality using the optimized media. One-sentence summary This study reports optimization of cell growth and expression conditions for a recombinant chimeric meningococcal protein in E. coli that assures its industrial scale production and suitability for preclinical and clinical studies as a vaccine component against Meningococcal B serogroup.

Background/Objective: Apart from the attributes such as cost, yields and consistency that define the feasibility of a manufacturing process, physicochemical and immunological quality traits equally signify the functionality of a biological product. The present study investigates one such promising Meningococcal B vaccine candidate, a chimeric fHbp-PorA protein in Escherichia coli. Methods: The chimeric fHbp–PorA protein, expressed with an N-terminal tag of HIS-MBP-TEV was produced in a 10 L fermenter under two different media and induction strategies: chemically defined (CD) media with lactose induction and complex media (CM) with galactose-mediated autoinduction. Comparative analysis was carried out between the two approaches for cell growth, protein expression, and purification, and the final chimeric proteins were characterized to evaluate for their biochemical, structural, in vitro and in vivo immunochemical properties. Results: Growth in the CD media resulted in several-fold-higher biomass compared to that in CM media in a short cultivation time; however, more than a third of the expressed protein remained in an insoluble state. Meanwhile, almost all of the expressed protein with CM media was recovered in soluble form. Moreover, purification of the unprocessed tagged protein and recovery of chimeric protein (tag removed) resulted in 75% greater yield in CM media when compared to CD media. The final chimeric proteins obtained from each medium varied significantly in their physicochemical characteristics, including their epitope projection and CD spectra. The results of in vivo animal immunogenicity response also showed higher serum bactericidal activity associated with chimeric protein obtained from CM media compared to CD media. Conclusions: The outcomes demonstrate that complex media with galactose-induced expression not only show higher productivity but also exhibit superior quality attributes, qualifying their reliable use in the manufacturing process of this promising vaccine candidate.