Expression and characterization of a novel PPO gene from Mucuna pruriens (L.) DC. var. pruriens involved in catecholamine pathway mediated synthesis of L-DOPA

• Published in: South African journal of botany 2023-07, Vol.158, p.232-242
• Main Authors: G, Saranya, M V, Jiby, Nair, R. Aswati, Pillai, Padmesh P., Jayabaskaran, C.
• Format: Article
• Language: English
• Subjects: L-DOPA; Mucuna pruriens; PPO; qRT- PCR; SDS-PAGE
• ISSN: 0254-6299; 1727-9321
• DOI: 10.1016/j.sajb.2023.04.040

Mucuna pruriens (L.) DC var. pruriens is a well-known legume of Fabaceae family. The plant is the natural source of l-DOPA, precursor molecule of dopamine neurotransmitter used in the treatment of Parkinson´s disease. In M. pruriens, the Catecholamine (CA) biosynthesis is mediated by Polyphenol oxidase (PPO) as validated in our previous reports. In the present investigation, we identified three PPO isoforms and were quantitatively analyzed by quantitative real-time PCR to understand the expression of PPO´s constitutively in the different tissues of M. pruriens. Consequently, the highly expressed PPO isoform was cloned, and the recombinant protein purified followed by functional characterization. The Kinetic analysis using Lineweaver-Burk curves showed greater binding affinity of enzyme towards its specific substrate. This study provides new insight into plant derived recombinant PPO enzyme, which is involved in CA biosynthetic pathway in this taxon. The Metabolic pathway engineering of PPO step would result in modulation of l-DOPA synthesis in this plant, in future. [Display omitted] •Identified and confirmed the existence of PPO as isoforms in M. pruriens.•Quantitative expression of the highest expressed isoform using qRT-PCR.•Functional characterization of recombinantly expressed purified PPO.