Liquid-liquid extraction of protease from cold-adapted yeast Rhodotorula mucilaginosa L7 using biocompatible and biodegradable aqueous two-phase systems

Author(s)

Lario, Luciana Daniela ^[1] ; Pellegrini Malpiedi, Luciana ^{[1, 2]} ; Brandao Pereira, Jorge Fernando ^[3] ; Sette, Lara Duraes ^[4] ; Pessoa-Junior, Adalberto ^[1]

Affiliation

^[1] Univ Sao Paulo, Dept Biochem & Pharmaceut Technol, Sao Paulo - Brazil

^[2] Univ Nacl Rosario, Fac Ciencias Bioquim & Farmaceut, Dept Quim Fis, RA-2000 Rosario, Santa Fe - Argentina

^[3] Univ Estadual Paulista, UNESP, Sch Pharmaceut Sci, Dept Bioproc & Biotechnol, Araraquara, SP - Brazil

^[4] Univ Sao Paulo UNESP, Inst Biosci, Dept Biochem & Microbiol, Sao Paulo - Brazil

• Document type: Journal article
• Source: SEPARATION SCIENCE AND TECHNOLOGY; v. 51, n. 1, p. 57-67, JAN 2 2016.
• Web of Science Citations: 6

Abstract

This work aimed to optimize the extraction of an extracellular protease produced by the cold-adapted yeast Rhodotorula mucilaginosa L7 using aqueous two-phase systems (ATPS) comprising polyethylene glycol (PEG) and sodium citrate or sodium tartrate. First, the biocompatibility of the phase forming agents was assessed. The results obtained with PEG-2000, PEG-4000, and PEG-6000 demonstrated that even at large PEG concentrations (32 wt%) the protease maintains its activity after 3 h of reaction, whereas an increase in salt concentration provokes a gradual decrease in protease stability. Subsequently, the partitioning of the protease in both types of ATPS was assessed, evaluating the effect of temperature, molecular weight, and concentration of PEG on protease purification, using two 2(3)-full factorial designs. The best partitioning conditions were obtained in PEG-6000/sodium tartrate-based ATPS, at 30oC (with a yield of 81.09 +/- 0.66% and a purification factor of 2.51 +/- 0.03). Thus, considering the biodegradable characteristics of the system, the PEG/sodium tartrate ATPS is a viable and economic low-resolution step in protease purification, with a strong potential for future industrial application. (AU)

• FAPESP's process: 13/19486-0 - Marine and Antarctic biotechnology: microbial enzymes and their applications
• Grantee: Lara Durães Sette
• Support Opportunities: Regular Research Grants
• FAPESP's process: 11/20521-0 - Extraction of monoclonal antibody fragments type scFv by aqueous two-phase micellar systems
• Grantee: Luciana Pellegrini Malpiedi
• Support Opportunities: Scholarships in Brazil - Post-Doctoral
• FAPESP's process: 12/23726-4 - "purification of proteases secreted by the psicrotroph yeast Rhodoturola mucilagionosa by aqueous two-phase systems (PEG/NaPA) and chromatographic processes"
• Grantee: Luciana Daniela Lario
• Support Opportunities: Scholarships in Brazil - Post-Doctoral