N-Glycosidase (PNGase) is a type of glycoside hydrolase whose main function is to release N-glycan chains from glycopeptides or glycoproteins, holding significant application value in fields such as food and biomedicine.Currently, commercial PNGases exhibit certain limitations in their application.Therefore, developing new PNGases with superior performance holds research value.This study successfully obtained a novel PNGase, named PNGase Dc, by expressing the gene encoding PNGase from Dyella caseinilytica in Escherichia coli.The molecular weight of PNGasse Dc was 63.01 kDa.Its enzymatic characteristics, functionality, and potential applications were then analyzed.The enzymatic characterization results indicated that the optimal pH and temperature for PNGase Dc are 2.0 and 37 ℃, respectively, and its catalytic reaction did not depend on metal ions.The enzyme also demonstrated excellent stability, retaining its activity even after storage at 4 ℃ for 45 days.Molecular docking analysis indicated that PNGase Dc interacts with its substrate N′N-diacetylchitobiose through hydrogen bonds and van der Waals forces.Site-directed mutagenesis identified Asp102 and Glu236 as key amino acids affecting the enzyme′s activity.PNGase Dc exhibited effective deglycosylation on standard glycoproteins, including horseradish peroxidase and lactoferrin.Lastly, PNGase Dc′s application potential was evaluated.Arabidopsis thaliana and mouse plasma was used as substrates, confirming its promising applicability in the study of glycoproteins from both animal and plant sources.In summary, this research introduced a new enzymatic tool, PNGase Dc, for the study of glycoproteins derived from animal and plant sources, thereby laying a theoretical foundation for the enzyme′s development and utilization.
WU Junying
,
WANG Ting
,
JOSEF Voglmeir
,
LIU Li
. Characteristics and functional analysis of novel N-glycosidase derived from Dyella caseinilytica[J]. Food and Fermentation Industries, 2024
, 50(14)
: 104
-110
.
DOI: 10.13995/j.cnki.11-1802/ts.039310
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