Peptide-Mediated Gene Transfer into Marine Purple Photosynthetic Bacteria

• Main Authors: Mieko Higuchi-Takeuchi, Takaaki Miyamoto, Choon Pin Foong, Mami Goto, Kumiko Morisaki, Keiji Numata
• Format: Article
• Language: English
• Published: MDPI AG 2020-11-01
• Series: International Journal of Molecular Sciences
• Subjects: transformation; plasmid DNA delivery; <i>Rhodovulum sulfidophilum</i>; cell penetrating peptide
• Online Access: https://www.mdpi.com/1422-0067/21/22/8625
• ISSN: 1661-6596; 1422-0067

Use of photosynthetic organisms is one of the sustainable ways to produce high-value products. Marine purple photosynthetic bacteria are one of the research focuses as microbial production hosts. Genetic transformation is indispensable as a biotechnology technique. However, only conjugation has been determined to be an applicable method for the transformation of marine purple photosynthetic bacteria so far. In this study, for the first time, a dual peptide-based transformation method combining cell penetrating peptide (CPP), cationic peptide and Tat-derived peptide (dTat-Sar-EED) (containing D-amino acids of Tat and endosomal escape domain (EED) connected by sarcosine linkers) successfully delivered plasmid DNA into <i>Rhodovulum sulfidophilum,</i> a marine purple photosynthetic bacterium. The plasmid delivery efficiency was greatly improved by dTat-Sar-EED. The concentrations of dTat-Sar-EED, cell growth stage and recovery duration affected the efficiency of plasmid DNA delivery. The delivery was inhibited at 4 °C and by A22, which is an inhibitor of the actin homolog MreB. This suggests that the plasmid DNA delivery occurred via MreB-mediated energy dependent process. Additionally, this peptide-mediated delivery method was also applicable for <i>E. coli </i>cells. Thus, a wide range of bacteria could be genetically transformed by using this novel peptide-based transformation method.