In silico structural characterization of the geranyl pyrophosphate synthase (GPPS) large subunit from Catharanthus...

Catharanthus roseus, a medicinal plant known for its anti-cancer monoterpene indole alkaloids vinblastine and vincristine, possesses two distinct types of geranyl pyrophosphate synthases (GPPS). A homomeric GPPS synthesizes geranyl pyrophosphate (GPP) as the sole product in the mitochondria, while a heteromeric GPPS comprising of a large subunit (LSU) and small subunit (SSU) is responsible for the GPP production in the chloroplasts. In the present study, in silico structural characterization and structure-activity correlation analysis of large subunit of the heteromeric GPPS (CrGPPS.LSU, GenBank Accession: JX417183) of C. roseus has been carried out. CrGPPS.LSU belongs to the type II trans-prenyltransferase family, having seven conserved regions. The protein comprises of 383 amino acids, with the first 63 amino acids belonging to the chloroplast transfer signal peptide sequence. CrGPPS.LSU showed the highest sequence identity with the heterotetrameric geranyl pyrophosphate synthase from Mentha piperita. A 3 dimensional (3D) structural model of the protein was obtained using AlphaFold 3, and was compared with other structurally characterized GPPS/GGPPS from plants, which revealed specific amino acid residues responsible for product formation in CrGPPS.LSU. The homomeric association of CrGPPS.LSU may be mediated through amino acid residues H109, D141, E149 and K172, while the presence of F148 might restrict the product chain elongation. An in silico mutation analysis of selected amino acids of CrGPPS.LSU further confirmed that the heterodimeric association of CrGPPS.LSU with a Type-I small subunit (CrGPPS.SSU; GenBank Accession: JX417184) is stabilized through interactions involving amino acid residues H109, D141, E133, D134, E149 and E177.