Structure-based analysis of Clot as a thioredoxin-related protein of 14 kDa in Drosophila via experimental and computational approaches.

The clot gene is required for the biosynthesis of drosopterins, the red components of Drosophila eye pigments. However, the enzymatic role of Clot in Drosophila eye pigment formation and the molecular mechanisms underlying Clot function are not fully elucidated. In this study, we cloned and characterized Clot derived from Drosophila cDNA, and results showed that Clot exhibited ∼30% sequence identity with mammalian TRP14. In addition, we reported the three-dimensional structure of Drosophila Clot based on homology modeling. Furthermore, we identified NFκB as a novel Clot substrate using the I-TASSER program. The NFκB fragment can bind near the active site of Clot. These findings predicted the novel regulatory mechanisms underlying Clot function in the pyrimidodiazepine synthesis pathway and increased the understanding of the molecular mechanisms and physiological function of Clot in Drosophila eye pigment formation.