Insects are the most diverse group of organisms on earth, colonizing almost every ecological niche of the planet. To survive in various and sometimes extreme habitats, insects have established diverse biological and chemical systems. Core components of these systems are enzymes that enable the insects to feed on diverse nutrient sources. The enzymes are produced by either the insects themselves (homologous) or by symbiotic organisms located in the insects' bodies or in their nests (heterologous). The use of these insect-associated enzymes for applications in the fields of food biotechnology and industrial (white) biotechnology is gaining more and more interest. Prominent examples of insect-derived enzymes include peptidases, amylases, lipases, and β-D-glucosidases. Highly potent peptidases for the degradation of gluten, a storage protein that can cause intestinal disorders, may be received from grain pests. Several insects, such as bark and ambrosia beetles and termites, are able to feed on wood. In the field of white biotechnology, their cellulolytic enzyme systems of mainly endo-1,4-β-D-glucanases and β-D-glucosidases can be employed for saccharification of the most prominent polymer on earth-cellulose.
Various food proteins including, e.g. gluten, collagen and casein are rich in L-proline residues. Due to the cyclic structure of proline, these proteins are well protected from enzymatic degradation by typical digestive enzymes. Proline-specific peptidases (PsP) belong to different families of hydrolases acting on peptide bonds (EC 3.4.x.x). They occur in various organisms including bacteria, fungi, plants and insects. Based on their biochemical characteristics, PsP type enzymes are further grouped into different subclasses of which prolyl aminopeptidases (EC 3.4.11.5, PAP), prolyl carboxypeptidases (EC 3.4.17.16, PCP) and prolyl oligopeptidases/prolyl endopeptidases (EC 3.4.21.26, POP/PEP) are of major interest for applications in food biotechnology. This mini review summarises the biochemical assays employed for these subclasses of PsP and their structural properties and the reaction mechanisms. A special focus was set on PsP derived from fungi and insects and important industrial applications in the field of food biotechnology. The degradation of gluten and collagen as well as the hydrolysis of bitter peptides are discussed.
As requirements for Advanced Therapy Medicinal Product (ATMP) production differ from other production processes (e.g., therapeutic protein production), cell detachment is often a crucial step for the process success. In most cases, cell detachment is done enzymatically. Although many peptidases are established in cell culture in R&D, e.g., Trypsin as gold standard, many of them seem to be unsuitable in ATMP production processes. Therefore, the present study investigated a novel endopeptidase used in food biotechnology for its applicability in ATMP processes where cell detachment is needed. The Prolyl-specific Peptidase (PsP) is of nonmammalian origin and considered as safe for humans. PsP was purified from the supernatant of the fungus Wolfiporia cocos. The isolation and purification resulted in an enzyme solution with 0.19 U mg −1 prolylspecific activity. By in silico analysis it was confirmed that attachment-promoting proteins can be cleaved by PsP in a similar amount than with Trypsin. Further the proteolytic activity was determined for PsP and Trypsin by using the same enzymatic assay. Detachment with both enzymes was compared for cells used in typical therapeutic production processes namely a mesenchymal stem cell line (hMSC-TERT) as a model for a cell therapeutic, Vero and MA104 cells used for viral therapeutic or vaccine production. The cell detachment experiments were performed with comparable enzyme activities (1.6 U mL −1). hMSC-TERT detachment was faster with PsP than with Trypsin. For Vero cells the detachment with PsP was not only faster but also more efficient. For MA104 cells the detachment rate with PsP was similar to Trypsin. For all cell types, detachment with PsP showed less influence on cell growth and metabolism compared to standard Trypsin.Thus, three cell types used in ATMP, viral therapeutics or vaccine production can be detached efficiently and gently with PsP. Therefore, PsP shows potential for cell detachment in ATMP and viral/vaccine production processes.
Häufigkeiten HA-assoziierter N-Glykane. Eine b-Propiolacton-Inaktivierung der Virusernte beeinflusste das N-Glykosylierungsmuster nicht. Aktuelle Studien in einem transgenen T-Zell-Rezeptor-Mausmodell zeigen, dass die HA N-Glykosylierung einen signifikanten Einfluss auf die Immunogenität haben kann.
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