Project description:Proteins, lipids, and carbohydrates from the harmful algal bloom (HAB)-causing organism Pyrodinium bahamense were characterized to obtain insights into the biochemical processes in this environmentally relevant dinoflagellate. Shotgun proteomics using label-free quantitation followed by proteome mapping using the P. bahamense transcriptome and translated protein databases of Marinovum algicola, Alexandrium sp., Cylindrospermopsis raciborskii, and Symbiodinium kawagutii for annotation enabled the characterization of the proteins in P. bahamense. The highest number of annotated hits were obtained from M. algicola and highlighted the contribution of microorganisms associated with P. bahamense. Proteins involved in dimethylsulfoniopropionate (DMSP) degradation such as propionyl CoA synthethase and acryloyl-CoA reductase were identified, suggesting the DMSP cleavage pathway as the preferred route in this dinoflagellate. Most of the annotated proteins were involved in amino acid biosynthesis and carbohydrate degradation and metabolism, indicating the active roles of these molecules in the vegetative stage of P. bahamense. This characterization provides baseline information on the cellular machinery and the molecular basis of the ecophysiology of P. bahamense.
Project description:Cytochrome b5 (b5) is a small accessory protein involved in several reactions. The interaction of b5 with its partner proteins, e.g., cytochrome P450 enzymes, has received much attention and has been the subject of numerous studies. In order to obtain further insights into protein-protein interactions of b5, we modified it with a green fluorophore, Alexa 488, which has a maleimide linker. The labeling was applied to both WT (without Cys) and the T70C mutant. This study revealed labeled sites in WT b5 and confirmed the Cys-selective labeling in the T70C mutant.