title: Process Integration for the Disruption of Candida guilliermondii Cultivated in Rice Straw Hydrolysate and Recovery of Glucose-6-Phosphate Dehydrogenase by Aqueous Two-Phase Systems.
abstract: Remaining cells of Candida guilliermondii cultivated in hemicellulose-based fermentation medium were used as intracellular protein source. Recovery of glucose-6-phosphate dehydrogenase (G6PD) was attained in conventional aqueous two-phase systems (ATPS) was compared with integrated process involving mechanical disruption of cells followed by ATPS. Influences of polyethylene glycol molar mass (M PEG) and tie line lengths (TLL) on purification factor (PF), yields in top (Y T ) and bottom (Y B ) phases and partition coefficient (K) were evaluated. First scheme resulted in 65.9 % enzyme yield and PF of 2.16 in salt-enriched phase with clarified homogenate (M PEG 1500 g mol(-1), TLL 40 %); Y B of 75.2 % and PF B of 2.9 with unclarified homogenate (M PEG 1000 g mol(-1), TLL 35 %). The highest PF value of integrated process was 2.26 in bottom phase (M PEG 1500 g mol(-1), TLL 40 %). In order to optimize this response, a quadratic model was predicted for the response PFB for process integration. Maximum response achieved was PFB = 3.3 (M PEG 1500 g mol(-1), TLL 40 %). Enzyme characterization showed G6P Michaelis-Menten constant (K M ) equal 0.07-0.05, NADP(+) K M 0.02-1.98 and optimum temperature 70 °C, before and after recovery. Overall, our data confirmed feasibility of disruption/extraction integration for single-step purification of intracellular proteins from remaining yeast cells.
authors.affiliation: Departamento de Biotecnologia, Escola de Engenharia de Lorena, Universidade de São Paulo, Estrada Municipal do Campinho s/n, 12602-810, Lorena, SP, Brazil.
authors.forename: Daniela B
authors.initials: DB
authors.lastname: Gurpilhares
authors.forename: Adalberto
authors.initials: A
authors.lastname: Pessoa
authors.forename: Inês C
authors.initials: IC
authors.lastname: Roberto
mesh_terms: D002175:Candida / Q000201:enzymology / Q000254:growth & development; D005656:Fungal Proteins / Q000096:biosynthesis / Q000737:chemistry / Q000302:isolation & purification; D005954:Glucosephosphate Dehydrogenase / Q000096:biosynthesis / Q000737:chemistry / Q000302:isolation & purification; D008956:Models, Chemical; D012275:Oryza / Q000737:chemistry*; D011134:Polysaccharides / Q000737:chemistry*
chemical_list: D005656:Fungal Proteins; D011134:Polysaccharides; C007916:hemicellulose; D005954:Glucosephosphate Dehydrogenase