Production of Chloroplast-Derived Antimicrobial Peptides

Introduction

Antimicrobial peptides (AMPs) are compounds with α-helical structures that exist in animals, plants, and microorganisms, and their role is to control normal microbial flora and fight pathogens. The rapid bactericidal activity of AMP makes it a promising candidate for therapeutic anti-infective drugs. AMP is especially effective in the treatment of complex skin and soft tissue infections. The emergence of antibiotic-resistant microorganisms and increasing attention to the use of antibiotics have led to the rapid development of AMPs, which have promising applications in the fields of medicine, food, animal husbandry, agriculture, and aquaculture. An ideal option for genetic engineering of crop plants for disease resistance through the chloroplast genome. The antimicrobial peptide MSI-99 is an analog of magainin 2, a synthetic cleavage peptide. Studies have shown that MSI-99 expression in transgenic chloroplasts is a cost-effective recombinant expression system with great therapeutic potential.

Fig. 1. Classification of antimicrobial peptides. (Huan Y, et al., 2020)

Solutions

Our engineers are passionate about engineering chloroplasts to produce therapeutic proteins for research to treat or prevent human or animal disease. Lifeasible is an expert in the field of recombinant protein research and development, with a variety of mature and comprehensive platforms and technologies, providing global customers with professional solutions for the production of chloroplast-derived antimicrobial peptides.

Our engineers sought to chloroplast engineering the magainin analog Myp30 in transgenic tobacco to confer disease resistance to plant fungal and bacterial pathogens. Here, we provide an expression system for the introduction of MSI-99 into tobacco chloroplast transformation for high levels of protection against bacterial and fungal pathogens. In addition, we developed transformation vectors with Retrocyclin-101 (RC101) and Protegrin-1 (PG1) coding sequences for expression in tobacco chloroplasts for use as therapeutics for bacterial or viral infections. Our solution process is roughly as follows:

(1) Designing a chloroplast transformation vector with MSI-99 coding sequence.
(2) The vector was transformed into tobacco chloroplasts by biolistic method.
(3) The transformed shoots were screened by PCR technology to confirm the integration of the transgenic cassette into the chloroplast genome.
(4) The 3P and 3M primers were used to detect the site-specific integration of the selectable marker gene (aadA) into the chloroplast genome.
(5) Southern blot analysis whether MSI-99 is stably integrated into the tobacco chloroplast genome.
(6) Western blot analysis of expression stability of MSI-99 protein.

Features of Our Solutions

• RC101 and PG1 retained their antibacterial activities when expressed in chloroplasts.
• High dose release at the site of infection is achieved by expressing MSI-99 from the chloroplast genome.
• Due to the high copy number associated with chloroplast expression, greater amounts of AMP can be synthesized.
• The accumulation of MSI-99 in transgenic chloroplasts did not affect the normal growth and development of transgenic plants.

The chloroplast-transformed antimicrobial peptides are safe and effective, and also confer disease resistance to plant fungal and bacterial pathogens. Lifeasible has extensive knowledge and experience in the development and production processes of the chloroplast-derived antimicrobial peptides. Our mission is to provide customers with comprehensive, reliable, professional solutions to accelerate your research. If you are interested in our solutions for the production of chloroplast-derived antimicrobial peptides, please contact us at any time.

References

1. DeGray G, Rajasekaran K, et al. (2001) Expression of an antimicrobial peptide via the chloroplast genome to control phytopathogenic bacteria and fungi. Plant Physiol. 127(3):852-862.
2. Huan Y, Kong Q, Mou H, et al. (2020) Antimicrobial peptides: classification, design, application and research progress in multiple fields[J]. Frontiers in microbiology. 2559.