One-step high efficiency separation of prolyl endopeptidase from Aspergillus niger and its application.

Prolyl endopeptidase from Aspergillus niger (An-PEP) is an enzyme that recognizes C-terminal peptide bonds of amino acid chains and cleaves them by hydrolysis. An aqueous two-phase system (ATPS) was used to separate An-PEP from fermentation broth. Through single factor experiments, the ATPS containing 16 % (w/w) PEG2000 and 15 % (w/w) (NH ₄ ) ₂ SO ₄ at pH 6.0 obtained the recovery of 79.74 ± 0.16 % and the purification coefficient of 7.64 ± 0.08. It was then used to produce soy protein isolate peptide (SPIP) by hydrolysis of soy protein isolate (SPI), and SPIP-Ferrous chelate (SPIP-Fe) was prepared with SPIP and Fe ²⁺ . The chelation conditions were optimized by RSM, as the chelation time was 30 min, chelation temperature was 25 °C, SPIP mass to VC mass was two to one and pH was 6.0. The obtained chelation rate was 82.56 ± 2.30 %. The change in the structures and functional features of SPIP before and after chelation were investigated. The FTIR and UV-Vis results indicated that the chelation of Fe ²⁺ and SPIP depended mainly on the formation of amide bonds. The fluorescence, SEM and amino acid composition analysis results indicated that Fe ²⁺ could induce and stabilize the surface conformation and change the amino acid distribution on the surfaces of SPIP. The chelation of SPIP and Fe ²⁺ resulted in the enhancement of radical scavenging activities and ACE inhibitory activities. This work provided a new perspective for the further development of peptide-Fe chelates for iron supplement.
Competing Interests: Declaration of competing interest The authors declare the following financial interests/personal relationships which may be considered as potential competing interests: Chunhong Liu reports financial support was provided by Heilongjiang Nature Scientific Foundation Project. If there are other authors, they declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.
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