title: The thermal unfolding of the ribosome-inactivating protein saporin-S6 characterized by infrared spectroscopy.
abstract: Saporin-S6 is a plant toxin belonging to the type 1 ribosome-inactivating protein (RIP) family. Since it was extracted and isolated from Saponaria officinalis for the first time almost thirty years ago, the protein has been widely studied mainly for its potential applications in anti-tumour and anti-viral infection therapy. Like other RIPs, saporin-S6 is particularly effective in the form of immunotoxin conjugated with monoclonal antibodies and its chemico-physical characteristics made the protein a perfect candidate for the synthesis, development and use of saporin-S6-based chimeric toxins. The high stability of the protein against different denaturing agents has been broadly demonstrated, however, its complete thermal unfolding characterization has not already been performed. In this work we analyse in detail structure, thermostability and unfolding features by means of infrared spectroscopy coupled with two-dimensional correlation spectroscopy. Our data showed that saporin-S6 in solution at neutral pH exhibits a secondary structure analogue to that of the crystal and confirmed its good stability at moderately high temperatures, with a temperature of melting of 58°C. Our results also demonstrated that the thermal unfolding process is non-cooperative and occurs in two steps, and revealed the sequence of the events that take place during the denaturation, showing a higher stability of the N-terminal domain of the protein.
authors.affiliation: Departamento de Bioquímica y Biología Molecular "A", Facultad de Veterinaria, Regional Campus of International Excellence "Campus Mare Nostrum", Universidad de Murcia, Murcia, Spain.
authors.forename: Marina
authors.initials: M
authors.lastname: Sánchez
authors.affiliation: Dipartimento di Scienze della Vita e dell'Ambiente, Università Politecnica delle Marche, Ancona, Italy.
authors.forename: Andrea
authors.initials: A
authors.lastname: Scirè
authors.affiliation: Dipartimento di Scienze della Vita e dell'Ambiente, Università Politecnica delle Marche, Ancona, Italy.
authors.forename: Fabio
authors.initials: F
authors.lastname: Tanfani
authors.affiliation: Prometeo Researcher, Departamento de Ciencias de la Salud, Universidad Técnica Particular de Loja (UTPL), Loja, Ecuador. Electronic address: aleausil@tin.it.
authors.forename: Alessio
authors.initials: A
authors.lastname: Ausili
mesh_terms: D006358:Hot Temperature; D006863:Hydrogen-Ion Concentration; D008958:Models, Molecular; D011489:Protein Denaturation; D055550:Protein Stability; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D058767:Protein Unfolding; D054789:Ribosome Inactivating Proteins, Type 1 / Q000737:chemistry* / Q000302:isolation & purification; D029750:Saponaria / Q000737:chemistry*; D000076985:Saporins; D012639:Seeds / Q000737:chemistry*; D012996:Solutions; D017550:Spectroscopy, Fourier Transform Infrared
chemical_list: D054789:Ribosome Inactivating Proteins, Type 1; D012996:Solutions; D000076985:Saporins
keywords: Infrared spectroscopy; Ribosome-inactivating protein; Saporin-S6; Thermal unfolding