Biochemical characterization, thermal stability, and partial sequence of a novel exo-polygalacturonase from the thermophilic fungus rhizomucor pusillus a13.36 obtained by submerged cultivation

• Autor(a) principal: Trindade, Lucas Vinícius [UNESP]
• Data de Publicação: 2016
• Outros Autores: Desagiacomo, Carla, Polizeli, Maria De Lourdes Teixeira De Moraes, Damasio, André Ricardo De Lima, Lima, Aline Margarete Furuyama [UNESP], Gomes, Eleni [UNESP], Bonilla-Rodriguez, Gustavo Orlando [UNESP]
• Tipo de documento: Artigo
• Idioma: eng
• Título da fonte: Repositório Institucional da UNESP
• Texto Completo: http://dx.doi.org/10.1155/2016/8653583; http://hdl.handle.net/11449/178483

This work reports the production of an exo-polygalacturonase (exo-PG) by Rhizomucor pusillus A13.36 in submerged cultivation (SmC) in a shaker at 45°C for 96 h. A single pectinase was found and purified in order to analyze its thermal stability, by salt precipitation and hydrophobic interaction chromatography. The pectinase has an estimated Mw of approximately 43.5-47 kDa and optimum pH of 4.0 but is stable in pH ranging from 3.5 to 9.5 and has an optimum temperature of 61°C. It presents thermal stability between 30 and 60°C, has 70% activation in the presence of Ca2+, and was tested using citrus pectin with a degree of methyl esterification (DE) of 26%. Ea(d) for irreversible denaturation was 125.5 kJ/mol with positive variations of entropy and enthalpy for that and ΔG(d) values were around 50 kJ/mol. The hydrolysis of polygalacturonate was analyzed by capillary electrophoresis which displayed a pattern of sequential hydrolysis (exo). The partial identification of the primary sequence was done by MS MALDI-TOF and a comparison with data banks showed the highest identity of the sequenced fragments of exo-PG from R. pusillus with an exo-pectinase from Aspergillus fumigatus. Pectin hydrolysis showed a sigmoidal curve for the Michaelis-Menten plot.