Articolo in rivista, 2018, ENG, 10.1007/s00425-018-2920-z

High-level production of single chain monellin mutants with enhanced sweetness and stability in tobacco chloroplasts

Daniela Castiglia, Serena Leone, Rachele Tamburino, Lorenza Sannino, Jole Fonderico, Chiara Melchiorre, Andrea Carpentieri, Stefania Grillo, Delia Picone, Nunzia Scotti

CNR-IBBR, Res. Div. Portici Dipartimento di Scienze Chimiche Università di Napoli Federico II

The prevalence of obesity and diabetes has dramatically increased the industrial demand for the development and use of alternatives to sugar and traditional sweeteners. Sweet proteins, such as MNEI, a single chain derivative of monellin, are the most promising candidates for industrial applications. In this work, we describe the use of tobacco chloroplasts as a stable plant expression platform to produce three MNEI protein mutants with improved taste profile and stability. All plant-based proteins were correctly expressed in tobacco chloroplasts, purified and subjected to in-depth chemical and sensory analyses. Recombinant MNEI mutants showed a protein yield ranging from 5% to more than 50% of total soluble proteins, which, to date, represents the highest accumulation level of MNEI mutants in plants. Comparative analyses demonstrated the high similarity, in terms of structure, stability and function, of the proteins produced in plant chloroplasts and bacteria. The high yield and the extreme sweetness perceived for the plant-derived proteins prove that plastid transformation technology is a safe, stable and cost-effective production platform for low-calorie sweeteners, with an estimated production of up to 25-30 mg of pure protein/plant.

Planta 248 , pp. 465–476

Keywords

Green factory, Plastid transformation, Nicotiana tabacum, Sweet proteins, Protein structure, Low-calorie sweeteners

CNR authors

Tamburino Rachele, Castiglia Daniela, Grillo Maria Stefania, Scotti Nunzia, Sannino Lorenza

CNR institutes

IBBR – Istituto di Bioscienze e Biorisorse