Immobilizationandpurification of soybean seed hull proteins of 9044 biotechnological interest identified by shotgun proteomic anal- 9045 ysis

LAUTARO BRACCO; F. WOLMAN; MARIA VICTORIA MIRANDA; CASCONE O

Congreso; European Federation of Biotechnology; 2018

Soybean crops have had a huge development. Soybean con- 9051sumption is led by oil and flour, for which the hull must be removed. 9052However, hull contains valuable proteins that can be extracted 9053prior to pellet it for animal food. The aim of this work was the 9054proteomic characterization of soybean seed hull aqueous extract 9055as a way to identify candidate proteins for downstream process- 9056ing, thus allowing the revaluation of this by-product of soybean 9057manufacture. A hull extract was concentrated by ultrafiltration and 9058subjected to analysis by nano-HPLC-ESI-Orbitrap. A total of 149 9059proteins were identified, among them 60 with potential biotech- 9060nological application, 15 in high abundance including peroxidase, 9061urease and glycinin. These proteins were immobilized on chitosan 9062mini-spheres. Peroxidase immobilization yield was over 90% when 9063mini-spheres were derivatized with the triazine dye Reactive Blue 90644, selected after a screening with 19 dyes. For urease and glycinin, 9065mini-spheres were chemically modified to obtain thiosulfonate derivatives, thus allowing the immobilization of urease (95% yield)9068 and the purification of glycinin by elutionwith -mercaptoethanol9069 (70% yield). For immobilized urease, the pH of maximal activity9070 shifted from 7 to 4?5, allowing its utilization in acidic media, i.e.9071 decrease of urea concentration in wine. Thermal, operational and9072 storage stability of the immobilized proteins were improved, as9073 comparedwith the free enzymes. Immobilized peroxidase and ure9074ase were applied to solve real problems: degradation of phenols in9075 wastewater and dosage of serum urea, respectively, along various9076 cycles with no significant decrease in enzyme activity.