The marine environment is a plentiful source of bioactive natural products that can assist us in the development of potential pharmaceutical agents. In the present study, we screened 300 bacteria from the Red Sea for their antitumor and antibacterial activities. The bioassay-guided analysis of the strain Streptomyces caelestis AW99C led to the discovery of two diastereoisomers citreamicin ε A and B, together with their derivatives. The antibacterial activities of all compounds were evaluated. Further cytotoxic mechanism study revealed that citreamicin ε could cause DNA fragmentation and induce caspase-3-dependent apoptosis in HeLa cells. Citreamicin ε greatly increased the intracellular ROS concentration in HeLa cells, suggesting the possible involvement of ROS in cell apoptosi...[ Read more ]

The marine environment is a plentiful source of bioactive natural products that can assist us in the development of potential pharmaceutical agents. In the present study, we screened 300 bacteria from the Red Sea for their antitumor and antibacterial activities. The bioassay-guided analysis of the strain Streptomyces caelestis AW99C led to the discovery of two diastereoisomers citreamicin ε A and B, together with their derivatives. The antibacterial activities of all compounds were evaluated. Further cytotoxic mechanism study revealed that citreamicin ε could cause DNA fragmentation and induce caspase-3-dependent apoptosis in HeLa cells. Citreamicin ε greatly increased the intracellular ROS concentration in HeLa cells, suggesting the possible involvement of ROS in cell apoptosis. Since citreamicin ε A and B had different apoptotic effects on PtK2 cells, we performed an iTRAQ based quantitative proteomic analysis in order to reveal the mechanism of cytotoxicity of citreamicin ε A and B in PtK2 cells. These significantly changed proteins were analyzed for their protein-protein interaction, which revealed the involvement of NF-κB pathway. Citreamicin ε A treatment induced rapid activation of the NF-κB pathway, which might have promoted cell survival and resulted in lower toxicity.

Biosynthetic studies in the present thesis refered to two kinds of natural products: one was polycyclic xanthone antibiotics and another one was hybrid polyketide-terpenoid halogenated meroterpenoids. The polycyclic xanthone antibiotics are synthesized by a type II polyketide synthase (PKS) and then undergo various post-PKS modifications to produce a variety of the polycyclic xanthone antibiotics. The entire 38-kb citreamicin (cit) gene cluster was sequenced, and in silica analysis revealed 36 open reading frames (ORFs) encoding type II polyketide synthases, regulators, transporters, and unique polyketide tailoring enzymes. Regarding to the marinones which are hybrid polyketide-terpenoid halogenated meroterpenoids, we adapted the transformation-associated recombination (TAR) to clone the targeted 61-kb marinone biosynthesis (mrn) gene cluster. In addition, the gene cluster unexpectedly harbored three vanadium-dependent haloperoxidases (VHPO) genes that putatively brominated the naphthoquinone core via novel bacterial halogenation biochemistry. To elucidate the divergent biosynthetic pathways of the marinones and the function of the haloperoxidases, we performed a series of in vivo and in vitro experiments. Our results confirmed that one of the VHPOs catalyzed the bromination of debromarinone and neomarinone.

The finding of this thesis showed that 1) the Red Sea microbes are potent and unexplored source of novel and bioactive compounds, 2) bioactive compounds indentified by me showed antitumor and antibacterial activities, 3) comparative proteomic analysis can be used to study the mode-of-action of stereoisomers in toxicology, and 4) the transformation-associated recombination cloning can be used in the biosynthetic study of natural products from microbes.