Characterization of a New Multifunctional GH20 β-N-Acetylglucosaminidase From Chitinibacter sp. GC72 and Its Application in Converting Chitin Into N-Acetyl Glucosamine

Abstract

In this study, a gene encoding β-N-acetylglucosaminidase, designated NAGaseA, was cloned from Chitinibacter sp. GC72 and subsequently functional expressed in Escherichia coli BL21 (DE3). NAGaseA contains a glycoside hydrolase family 20 catalytic domain that shows low identity with the corresponding domain of the well-characterized NAGases. The recombinant NAGaseA had a molecular mass of 92 kDa. Biochemical characterization of the purified NAGaseA revealed that the optimal reaction condition was at 40°C and pH 6.5, and exhibited great pH stability in the range of pH 6.5–9.5. The Vmax, Km, kcat, and kcat/Km of NAGaseA toward p-nitrophenyl-N-acetyl glucosaminide (pNP-GlcNAc) were 3333.33 μmol min–1 l–1, 39.99 μmol l–1, 4667.07 s–1, and 116.71 ml μmol–1 s–1, respectively. Analysis of the hydrolysis products of N-acetyl chitin oligosaccharides (N-Acetyl COSs) indicated that NAGaseA was capable of converting N-acetyl COSs ((GlcNAc)2–(GlcNAc)6) into GlcNAc with hydrolysis ability order: (GlcNAc)2 > (GlcNAc)3 > (GlcNAc)4 > (GlcNAc)5 > (GlcNAc)6. Moreover, NAGaseA could generate (GlcNAc)3–(GlcNAc)6 from (GlcNAc)2–(GlcNAc)5, respectively. These results showed that NAGaseA is a multifunctional NAGase with transglycosylation activity. In addition, significantly synergistic action was observed between NAGaseA and other sources of chitinases during hydrolysis of colloid chitin. Finally, 0.759, 0.481, and 0.986 g/l of GlcNAc with a purity of 96% were obtained using three different chitinase combinations, which were 1.61-, 2.36-, and 2.69-fold that of the GlcNAc production using the single chitinase. This observation indicated that NAGaseA could be a potential candidate enzyme in commercial GlcNAc production.

Analysis of Online and Offline Teaching Pathways for Aquatic Microbiology Based on OBE Concept

“…The closer the results are to the real learning experience of students, the more likely they are to persist, especially after long-term practice [2] .…”

Gene functions of the Ambystoma altamirani skin microbiome vary across space and time but potential antifungal genes are widespread and prevalent

“…Moreover, it has been shown that amphibians exposed to Bd exhibit an increased abundance of a bacterial-derived chitin deacetylase [ 22 ], an enzyme that contributes to chitin degradation [ 123 ]. Here we reported the presence of genes linked to chitin degradation in Burkholderiales and Chitinophagales MAGs including glycoside hydrolases from the GH19 [ 124 ], GH20 [ 125, 126 ], and GH23 [ 117, 127, 128 ] families. Noteworthy, GH23 enzymes were mainly identified in Burkholderiales MAGs, while GH20 enzymes were identified only in Chitinophagales MAGs suggesting these taxa may use different/complementary strategies for chitin degradation.…”

Catalytic conversion of chitin-based biomass to nitrogen-containing chemicals

“…Chitinases have been successfully extracted from various bacterial strains, such as Serratia proteamaculans NJ303, Aeromonas hydrophila NJ303, Chitinolyticbacter meiyuanensis SYBC-H1 and Chitinibacter sp. GC72, for the depolymerization of chitin, 77 , 78 , 79 , 80 highlighting the availability of microbial technology in sustainable NAG production. To improve efficiency, it is necessary to further develop the chitinases separation and purification technologies.…”