Molecular identification, isolation and functional characterization of a glutathione S-transferase gene CsGST in saffron (Crocus sativus L.)

Crocus sativus L., an herbaceous species belonging to the Iridaceae family is renowned for its medicinal properties. The vibrant color of its petals and leaves is attributed to anthocyanins, whereas the highly valued stigma predominantly accumulates apocarotenoids, such as crocin, as its main pigments. Glutathione S-transferases (GSTs) are multifunctional enzymes, with many members playing a crucial role in the transport of anthocyanins and other flavonoids. In this study, we isolated the cDNA sequence of a GST gene from saffron, designated CsGST, using RT-PCR and RACE techniques. The sequence revealed a 795 bp open reading frame encoding 264 amino acids. The corresponding genomic sequence was 1074 bp, containing two exons and one intron. Phylogenetic and conserved domain analysis classified CsGST as a Tau-type GST. Expression analysis detected CsGST transcripts in leaves, petals, pistils, and stamens, with its expression in petals showing a significant positive correlation with anthocyanin content. The recombinant CsGST protein was expressed in E. coli and exhibited fundamental GST enzymatic activity. Furthermore, molecular docking analysis suggested a potential interaction between CsGST and crocin, the main carotenoid pigment in the stigma. This study represents the first comprehensive identification and characterization of a GST gene in saffron, providing a foundational basis for future research into its roles in the transport of secondary metabolites.