Characterization of Fatty Acid Amide Hydrolases in Tomato
N-acylethanolamines(NAEs) are fatty acid amides derived from a minor membrane lipid constituent Nacylphosphatidylethanolamine, structurally consisting the linkage of fatty acid tothe ethanolamines. NAE is hydrolysed by fatty acid amide hydrolase (FAAH) into free fatty acid and ethanolamine in both plants and animals. In plants, FAAH gene has been thus far characterized in Arabidopsis, where it was shown to act as a modulator of endogenous NAE levels, seedling growth and their ability to respond to biotic and biotic stress. Based on the evidence that NAEs occur in tomato (Solanum lycopersicum) seeds, we hypothesized that there is a functional FAAH that hydrolyzes NAEs in tomato. To test this, we performed in silico analysis using AtFAAH sequence as a template and identified six orthologs in tomato. These six S. lycopersicum FAAH homologs have the characteristic amidase signature sequence and conserved catalytic residues. Protein structures of putative SlFAAH1 and 2 were predicted using molecular visualization system (PyMOL). They showed similar domain structure with minor differences in spatial arrangement when compared with that of AtFAAH. Among the six homologs only putative SlFAAH1 and SlFAAH2 expression levels were associated with seedling development. Therefore the study was focused on cloning and characterization of SlFAAH1 and 2. Thus far, full-length coding sequence of putative SlFAAH1 was cloned into a heterologous expression system and its expression was confirmed by Western blot. Biochemical characterization of its hydrolytic activity against radiolabelled NAE substrates is underway. Furthermore, expression of SlFAAH1 and SlFAAH2 will be quantified and correlated with the NAE levels and hydrolytic activity at different developmental stages. This study is expected to reveal how NAE metabolite levels are modulated in tomato plant during its development.
Johnson City, TN
Tiwari, Vijay; Stuffle, Derek; and Kilaru, Aruna. (false). 2016. Characterization of Fatty Acid Amide Hydrolases in Tomato. Appalachian Student Research Forum, Johnson City, TN. https://www.etsu.edu/studentresearch/2016/documents/2016_asrf_programbook.pdf