Abstract
Chlamydia is a prevalent sexually transmitted disease that infects more than 100 million people worldwide. Although most individuals infected with Chlamydia trachomatis are initially asymptomatic, symptoms can arise if left undiagnosed. Long-term infection can result in debilitating conditions such as pelvic inflammatory disease, infertility, and blindness. Chlamydia infection, therefore, constitutes a significant public health threat, underscoring the need for a Chlamydia-specific vaccine. Chlamydia strains express a major outer-membrane protein (MOMP) that has been shown to be an effective vaccine antigen. However, approaches to produce a functional recombinant MOMP protein for vaccine development are limited by poor solubility, low yield, and protein misfolding. Here, we used an Escherichia coli-based cell-free system to express a MOMP protein from the mouse-specific species Chlamydia muridarum (MoPn-MOMP or mMOMP). The codon-optimized mMOMP gene was co-translated with Δ49apolipoprotein A1 (Δ49ApoA1), a truncated version of mouse ApoA1 in which the N-terminal 49 amino acids were removed. This co-translation process produced mMOMP supported within a telodendrimer nanolipoprotein particle (mMOMP-tNLP). The cell-free expressed mMOMP-tNLPs contain mMOMP multimers similar to the native MOMP protein. This cell-free process produced on average 1.5 mg of purified, water-soluble mMOMP-tNLP complex in a 1-ml cell-free reaction. The mMOMP-tNLP particle also accommodated the co-localization of CpG oligodeoxynucleotide 1826, a single-stranded synthetic DNA adjuvant, eliciting an enhanced humoral immune response in vaccinated mice. Using our mMOMP-tNLP formulation, we demonstrate a unique approach to solubilizing and administering membrane-bound proteins for future vaccine development. This method can be applied to other previously difficult-to-obtain antigens while maintaining full functionality and immunogenicity.
Highlights
We used a cell-free method coupled with telodendrimer nanolipoprotein particles (tNLPs) technology to produce a recombinant mMOMP that is soluble and functional
We demonstrated that immunogenic adjuvants like CpG were incorporated into our mMOMP–tNLP formulation
Sera from mice immunized with mMOMP–tNLPs showed strong antibody titers that crossreacted with Chlamydia elementary body (EB)
Summary
As a funding agency they had no role in study design or data collection and analysis for publication of the data They do have a financial interest through the development of novel synthetic biology strategies for vaccine development. Using our mMOMP–tNLP formulation, we demonstrate a unique approach to solubilizing and administering membranebound proteins for future vaccine development This method can be applied to other previously difficult-to-obtain antigens while maintaining full functionality and immunogenicity. Their life cycles begin with a metabolically inactive infectious form called the elementary body (EB) that measures ϳ300 nm in diameter. Cell-free production of functional and immunogenic MoPn-MOMP severity of Chlamydia infection, even when asymptomatic, highlights the need for a long-term protective vaccine
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