Efficient Robust Yield Method for Preparing Bacterial Ghosts by Escherichia coli Phage ID52 Lysis Protein E

Abstract

Bacterial ghosts (BGs) are nonliving empty bacterial shells without cytoplasm retaining original morphology and identical antigenicity of natural bacteria, making them high potential and promising vaccine candidates and delivery vehicles. However, the low yield of commonly used BGs preparation methods limits its mass production and widely application. In order to improve BGs production, E. coli phage ID52 lysis protein E was introduced to generating BGs for the first time. Above all, we compared the lysis activity of lysis protein of E. coli phage φX174 and E. coli phage ID52 as well as the effects of promoters on the lysis activity of ID52-E, which shown that the lysis activity and BGs formation rate of protein ID52-E was significantly higher than protein φX174-E. Further, the lysis activity of ID52-E was significantly improved under the control of L-arabinose inducible promoter which initial induction OD600 reached as high as 2.0. The applicability of lysis protein ID52-E induced by L-arabinose was proved by preparing probiotic E. coli Nissle 1917 BGs and pathogenic Salmonella typhimurium BGs in mass production. This paper introduced a novel and highly efficient method for BGs preparation depending on recombinant expression of E. coli phage ID52-E under eco-friendly and reasonable price inducer L-arabinose.

Comparison of the immune effects of the Chlamydia abortus MOMP antigen displayed in different parts of bacterial ghosts

“…Referring to a previous study ( Jechlinger et al, 2005a ), the “WK-E” lysing cassette promoter was point-mutated ( Figure 1B ). The most visual method to assess the lytic activity of E protein was to measure the extent to which the OD 600 value decreased ( Ma et al, 2022 ). Therefore, the lysis activity of the pUC-ΔWK-E plasmid was evaluated in E. coli BL21 and pathogenic E. coli F107/86.…”

“…However, for the widespread adoption of BG vectors for use in vaccine development, it is necessary to develop a more convenient and efficient BG preparation system and simultaneously explore different strategies to enhance the capacity of BG vaccines to induce more efficient immune responses. In this study, we controlled E gene expression using the temperature-inducible expression plasmid pBV220 without adding other chemicals ( Ma et al, 2022 ). E gene expression is typically strictly suppressed at 28°C.…”

Construction and Mechanism Exploration of Highly Efficient System for Bacterial Ghosts Preparation Based on Engineered Phage ID52 Lysis Protein E

“…In recent years, the challenge of enhancing BG production has posed a significant concern among scholars, with BGs yet to receive official approval and commercialization [23]. There is a growing demand for high-efficiency BGs with increased yield and productivity, and as of now, no large-scale fermentation experiments have been conducted with EcN BGs.…”

“…However, the inducer isopropyl β-D-1-thiogalactopyranoside (IPTG) exhibits specific cytotoxic properties [22] and is cost-prohibitive, limiting its large-scale production and application in BGs. As an alternative, some researchers have recombinantly expressed the E. coli phage ID52 lysis protein ID52-E, utilizing an L-arabinose inducible promoter (araC-ParaBAD) to enhance the lysis effect of ID52-E [23], achieving an induction OD 600 as high as 2.5.…”

Production and Evaluation of the Immunogenicity of an Enterobacter aerogenes Ghost Vaccine in a Mouse Model

“…BGs are empty cell envelopes derived from gram-negative Egyptian Journal of Chemistry http://ejchem.journals.ekb.eg/ bacteria [9]. BGs preparation involves causing lysis of the bacterial cells through the controlled expression of the PhiX174 lysis gene E [10,11], or through the use of a minimum inhibitory concentration of certain chemicals [12,13]. BGs produced using the second method can be used to lyse not only gram-positive and gram-negative bacteria, but also yeast and fungal cells [14,15].…”