Biochemical Characterization and Structural Analysis of a β-N-Acetylglucosaminidase from Paenibacillus barengoltzii for Efficient Production of N-Acetyl-d-glucosamine

Abstract

Bioproduction of N-acetyl-d-glucosamine (GlcNAc) from chitin, the second most abundant natural renewable polymer on earth, is of great value in which chitinolytic enzymes play key roles. In this study, a novel glycoside hydrolase family-18 β-N-acetylglucosaminidase (PbNag39) from Paenibacillus barengoltzii suitable for GlcNAc production was identified and biochemically characterized. It possessed a unique shallow catalytic groove (5.8 Å) as well as a smaller C-terminal domain (solvent-accessible surface area, 5.1 × 103 Å2) and exhibited strict substrate specificity toward N-acetyl chitooligosaccharides (COS) with GlcNAc as the sole product, showing a typical manner of action of β-N-acetylglucosaminidases. Thus, an environmentally friendly bioprocess for GlcNAc production from ball-milled powdery chitin by an enzyme cocktail reaction was further developed. By using the new route, the powdery chitin conversion rate increased from 23.3% (v/v) to 75.3% with a final GlcNAc content of 22.6 mg mL–1. The efficient and environmentally friendly bioprocess may have great application potential in GlcNAc production.

Property and Function of a Novel Chitinase Containing Dual Catalytic Domains Capable of Converting Chitin Into N-Acetyl-D-Glucosamine

“…For (GlcNAc) 2–6 , Cm Chi3 slowly hydrolyzed (GlcNAc) 2 into GlcNAc, showing that Cm Chi3 possesses NAGase activity. This result is similar to that of other studies ( Liu Y. et al, 2020 ). However, (GlcNAc) 2 , (GlcNAc) 2–3 , (GlcNAc) 2–4 , and (GlcNAc) 2–5 were each released from (GlcNAc) 3 , (GlcNAc) 4 , (GlcNAc) 5 , and (GlcNAc) 6 , respectively, which suggested that Cm Chi3 has endocleavage activities.…”

Production of Lacto-N-triose II and Lacto-N-neotetraose from Chitin by a Novel β-N-Acetylhexosaminidase Expressed in Pichia pastoris

“…Chitin was pretreated by a CJM-SY-B vibration ball milling (Taiji Ring Nano, China) based on the method characterized in our previous research . 100 g of powdery chitin samples were mixed with 200 g of zirconia beads in zirconia canister for 4 h. The rotation speed was set at 380 rpm.…”

Heterogenously‐expressed chitosanase combining a green ball milling method for enzymatic degradation

“…The particle size of the BMPC reduced dramatically after pretreatment by ballmilling (Figure 6b), suggesting the structure destruction of PC during the severe physical friction (Schneider et al, 2017). The overall spectra of PC and BMPC were identical with the same characteristic peaks, indicating that ball-milling pretreatment did not affect the bonds between chitosan molecules to change (Liu et al, 2020) (Figure S2). As shown in Figure 7, the XRD spectra of PC exhibited two typical diffraction peaks with different intensities at roughly 2θ of 10.4°and 20.0°, implying that hydrogen interactions between amino and hydroxyl groups in PC induce crystallinity.…”