Cell-free production of a functional oligomeric form of a Chlamydia major outer-membrane protein (MOMP) for vaccine development

• DOI: 10.1074/jbc.m117.784561
• Publication Date: 2017-07-25T00:20:13Z
• Authors: Wei He; Martina Felderman; Angela C. Evans; Jia Geng; David Homan; Feliza Bourguet; Nicholas O. Fischer; Yuanpei Li; Kit S. Lam; Aleksandr Noy; Li Xing; R. Holland Cheng; Amy Rasley; Craig D. Blanchette; Kurt Kamrud; Nathaniel Wang; Heather Gouvis; Todd C. Peterson; Bolyn Hubby; Matthew A. Coleman
• Keywords: 0301 basic medicine; 570; Biochemistry & Molecular Biology; Chlamydia muridarum; Biomedical and clinical sciences; Clinical Sciences; apolipoprotein; Medical and Health Sciences; oligomer; Vaccine Related; Mice; 03 medical and health sciences; Animals; membrane protein; Chlamydia; Inbred BALB C; nanolipoproteins; Mice, Inbred BALB C; Biomedical and Clinical Sciences; nanotechnology; Base Sequence; Cell-Free System; Prevention; cell-free expression; major outer membrane protein; Biological Sciences; Chlamydia Infections; telodendrimer; 3. Good health; Biological sciences; Infectious Diseases; Good Health and Well Being; 3.4 Vaccines; Chemical sciences; Medical Microbiology; Chemical Sciences; Bacterial Vaccines; Sexually Transmitted Infections; Immunization; Female; Infection; Biotechnology; Bacterial Outer Membrane Proteins

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.