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BackgroundN-acetyl-β-D-glucosamine (GlcNAc) is widely used as a valuable pharmacological agent and a functional food additive. The traditional chemical process for GlcNAc production has some problems such as high production cost, low yield, and acidic pollution. Hence, to identify a novel chitinase that is suitable for bioconversion of chitin to GlcNAc is of great value.ResultsA novel chitinase gene (PbChi74) from Paenibacillus barengoltzii was cloned and heterologously expressed in Escherichia coli as an intracellular soluble protein. The gene has an open reading frame (ORF) of 2,163 bp encoding 720 amino acids. The recombinant chitinase (PbChi74) was purified to apparent homogeneity with a purification fold of 2.2 and a recovery yield of 57.9%. The molecular mass of the purified enzyme was estimated to be 74.6 kDa and 74.3 kDa by SDS-PAGE and gel filtration, respectively. PbChi74 displayed an acidic pH optimum of 4.5 and a temperature optimum of 65°C. The enzyme showed high activity toward colloidal chitin, glycol chitin, N-acetyl chitooligosaccharides, and p-nitrophenyl N-acetyl β-glucosaminide. PbChi74 hydrolyzed colloidal chitin to yield N-acetyl chitobiose [(GlcNAc)2] at the initial stage, which was further converted to its monomer N-acetyl glucosamine (GlcNAc), suggesting that it is an exochitinase with β-N-acetylglucosaminidase activity. The purified PbChi74 coupled with RmNAG (β-N-acetylglucosaminidase from Rhizomucor miehei) was used to convert colloidal chitin to GlcNAc, and GlcNAc was the sole end product at a concentration of 27.8 mg mL-1 with a conversion yield of 92.6%. These results suggest that PbChi74 may have great potential in chitin conversion.ConclusionsThe excellent thermostability and hydrolytic properties may give the exochitinase great potential in GlcNAc production from chitin. This is the first report on an exochitinase with N-acetyl-β-D-glucosaminidase activity from Paenibacillus species.

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A novel β-N-acetylglucosaminidase gene (RmNag) from Rhizomucor miehei was cloned and expressed in Escherichia coli. RmNag shares the highest identity of 37% with a putative β-N-acetylglucosaminidase from Aspergillus clavatus. The recombinant enzyme was purified to homogeneity. The optimal pH and temperature of RmNag were pH 6.5 and 50 °C, respectively. It was stable in the pH range 6.0-8.0 and at temperatures below 45 °C. RmNag exhibited strict substrate specificity for p-nitrophenyl β-N-acetylglucosaminide (pNP-GlcNAc) and N-acetyl chitooligosaccharides. The apparent Km of RmNag toward pNP-GlcNAc was 0.13 mM. The purified enzyme displayed an exo-type manner as it released the only end product of GlcNAc from all the tested N-acetyl chitooligosaccharides. Besides, RmNag exhibited relatively high N-acetyl-β-D-glucosaminide tolerance with an inhibition constant Ki value of 9.68 mM. The excellent properties may give the enzyme great potential in industries. This is the first report on a glycoside hydrolyase family 3 β-N-acetylglucosaminidase from a fungus.

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Let (X , d, µ) be a metric measure space satisfying both the upper doubling and the geometrically doubling conditions. In this paper, the authors establish some equivalent characterizations for the boundedness of fractional integrals over (X , d, µ). The authors also prove that multilinear commutators of fractional integrals with RBMO(µ) functions are bounded on Orlicz spaces over (X , d, µ), which include Lebesgue spaces as special cases. The weak type endpoint estimates for multilinear commutators of fractional integrals with functions in the Orlicz-type space Osc exp L r (µ), where r ∈ [1, ∞), are also presented. Finally, all these results are applied to a specific example of fractional integrals over non-homogeneous metric measure spaces.

Let (X , d, µ) be a space of homogeneous type in the sense of Coifman and Weiss. In this article, the authors establish a complete real-variable theory of Musielak-Orlicz Hardy spaces on (X , d, µ). To be precise, the authors first introduce the atomic Musielak-Orlicz Hardy space H ϕ at (X) and then establish its various maximal function characterizations. The authors also investigate the Littlewood-Paley characterizations of H ϕ at (X) via Lusin area functions, Littlewood-Paley g-functions and Littlewood-Paley g * λ-functions. The authors further obtain the finite atomic characterization of H ϕ at (X) and its improved version in case q < ∞, and their applications to criteria of the boundedness of sublinear operators from H ϕ at (X) to a quasi-Banach space, which are also applied to the boundedness of Calderón-Zygmund operators. Moreover, the authors find the dual space of H ϕ at (X), namely, the Musielak-Orlicz BMO space BMO ϕ (X), present its several equivalent characterizations, and apply it to establish a new characterization of the set of pointwise multipliers for the space BMO(X). The main novelty of this article is that, throughout the article, except the last section, µ is not assumed to satisfy the reverse doubling condition.

CAS(14,12)/cc-pvdz calculations are reported for the reaction of 3CH2+3O2→products. On the singlet potential energy surface, a transition state has been located with an energy barrier of 1.65 kcal/mol, which is in good agreement with the experimental estimation of 1.0−1.5 kcal/mol. The rearrangement and metathesis of the singlet intermediates have been also investigated at the same level of theory. For the triplet case, the formation of CH2OO has an energy barrier of 5.79 kcal/mol, and the formed triplet CH2OO could be further decomposed into CH2O+O(3P) with an energy barrier of 2.92 kcal/mol. The geometries of some key points have been relocated at the CAS(8,6)+1+2/cc-pvdz level of theory for comparison. The present theoretical results for the total reaction rates, at the CAS(8,6)+1+2/cc-pvdz level, can be expressed by the three-parameter expression:  k(T) = 4.273×10-18 T 2.245exp(−185/T) within ±5% error at the temperature range 295−2600 K.