Genetic analysis, purification and docking studies of trihydroxynaphthalene reductase involved in pathogenesis of rice pathogen, Curvularia lunata

Abstract

Curvularia lunata causes black kernel disease in rice (Oryza sativa) kernel and enzyme, trihydroxynaphthalene reductase (3HNR) catalyzes a crucial reaction in melanin biosynthesis which aids in appressorium formation. However, the molecular mechanism of pathogenesis and characterization of 3HNR are poorly understood due to the limited research conducted on this pathosystem. Therefore, the C. lunata (GVBR1) was isolated from the diseased rice kernel and identified using microscopic, SEM and molecular methods. To elucidate the molecular mechanism of 3HNR, the C. lunata was treated with different concentrations of systemic fungicide tricyclazole and the gene expression of C. lunata trihydroxynaphthalene reductase (Cl3HNR) analyzed with different concentrations of tricyclazole treated, downregulated the Cl3HNR gene expression as compared to the control. Further, Cl3HNR was cloned into pGEX expression vector, heterologously expressed in Escherichia coli BL21 cells and the expressed protein corresponds to 28 kDa. The molecular docking and dynamic simulation revealed the interaction points within the active site of Cl3HNR. This study highlights the importance of the melanin biosynthetic gene involved in the pathogenesis of the rice fungal pathogen, C. lunata, which paves the way in developing the HNR enzyme inhibitor to control the rice disease.