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

Autor(es)

Gurpilhares, Daniela B. ^{[1, 2]} ; Pessoa, Adalberto ^[3] ; Roberto, Ines C. ^[1]

Afiliação do(s) autor(es)

^[1] Univ Sao Paulo, Escola Engn Lorena, Dept Biotecnol, BR-12602810 Lorena, SP - Brazil

^[2] Univ Fed Rio de Janeiro, Fac Farm, BR-27930560 Macae, RJ - Brazil

^[3] Univ Sao Paulo, Sch Pharmaceut Sci, Dept Biochem & Pharmaceut Technol, BR-05508900 Sao Paulo, SP - Brazil

• Tipo de documento: Artigo Científico
• Fonte: Applied Biochemistry and Biotechnology; v. 176, n. 6, p. 1596-1612, JUL 2015.
• Citações Web of Science: 0

Resumo

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 PFB 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 degrees 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. (AU)

• Processo FAPESP: 03/12855-9 - Purificação de enzima glicose-6-fosfato desidrogenase por processo de extração líquido-líquido em sistemas aquosos bifásicos integrado ao rompimento celular de Candida guilliermondii
• Beneficiário: Daniela de Borba Gurpilhares
• Linha de fomento: Bolsas no Brasil - Doutorado