Transglycosylation by a chitinase from Enterobacter cloacae subsp. cloacae generates longer chitin oligosaccharides

Mallakuntla, Mohan Krishna; Vaikuntapu, Papa Rao; Bhuvanachandra, Bhoopal; Das, Subha Narayan; Podile, Appa Rao

doi:10.1038/s41598-017-05140-3

Cited by 39 publications

(21 citation statements)

References 61 publications

(87 reference statements)

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“…The hydrolysis profile of Cm Chi1 is different from that of most chitinases from T. kodakaraensis KOD1 [ 42 ], Enterobacter cloacae subsp. cloacae [ 43 ] and P. barengoltzii [ 12 ]. For instance, Fu et al reported an exochitinase Pb Chi74 from P. barengoltzii possesses both exo activity and N -acetyl-β- d -glucosaminidase activity but no endo activity [ 12 ].…”

Section: Resultsmentioning

confidence: 99%

Molecular characterization of a novel chitinase CmChi1 from Chitinolyticbacter meiyuanensis SYBC-H1 and its use in N-acetyl-d-glucosamine production

Zhang

Wei

et al. 2018

Biotechnol Biofuels

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BackgroundN-acetyl-d-glucosamine (GlcNAc) possesses many bioactivities that have been used widely in many fields. The enzymatic production of GlcNAc is eco-friendly, with high yields and a mild production process compared with the traditional chemical process. Therefore, it is crucial to discover a better chitinase for GlcNAc production from chitin.ResultsA novel chitinase gene (Cmchi1) cloned from Chitinolyticbacter meiyuanensis SYBC-H1 and expressed in Escherichia coli BL21(DE3) cells. The recombinant enzyme (CmChi1) contains a glycosyl hydrolase family 18 catalytic module that shows low identity (12–27%) with the corresponding domain of the well-characterized chitinases. CmChi1 was purified with a recovery yield of 89% by colloidal chitin affinity chromatography, whereupon it had a specific activity of up to 15.3 U/mg. CmChi1 had an approximate molecular mass of 70 kDa after the sodium dodecyl sulfate-polyacrylamide gel electrophoresis, and its optimum activity for colloidal chitin (CC) hydrolysis occurred at pH 5.2 and 50 °C. Furthermore, CmChi1 exhibited kcat/Km values of 7.8 ± 0.11 mL/s/mg and 239.1 ± 2.6 mL/s/μmol toward CC and 4-nitrophenol N,N′-diacetyl-β-d-chitobioside [p-NP-(GlcNAc)2], respectively. Analysis of the hydrolysis products revealed that CmChi1 exhibits exo-acting, endo-acting and N-acetyl-β-d-glucosaminidase activities toward N-acetyl chitooligosaccharides (N-acetyl CHOS) and CC substrates, behavior that makes it different from typical reported chitinases. As a result, GlcNAc could be produced by hydrolyzing CC using recombinant CmChi1 alone with a yield of nearly 100% and separated simply from the hydrolysate with a high purity of 98%.ConclusionThe hydrolytic properties and good environmental adaptions indicate that CmChi1 has excellent potential in commercial GlcNAc production. This is the first report on exo-acting, endo-acting and N-acetyl-β-d-glucosaminidase activities from Chitinolyticbacter species.Electronic supplementary materialThe online version of this article (10.1186/s13068-018-1169-x) contains supplementary material, which is available to authorized users.

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Section: Resultsmentioning

confidence: 99%

Molecular characterization of a novel chitinase CmChi1 from Chitinolyticbacter meiyuanensis SYBC-H1 and its use in N-acetyl-d-glucosamine production

Zhang

Wei

et al. 2018

Biotechnol Biofuels

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“…Hydrolysis and transglycosylation (TG) are two significant processes that may determine the suitability of chitinases for specific roles. Chitinases interact with chitin either hydrolytically or by TG and some uniquely exhibit both properties [ 84 ]. Hydrolysis ( Figure 8 C) results in the generation of monomeric or dimeric glucosamine units with low degree of polymerization (DP) [ 24 ].…”

Section: Classification Based On Differences In Catalytic Mechanismsmentioning

confidence: 99%

“…Serratia proteamaculans ChiD ( Sp ChiD) was found to exhibit ‘transglyco-hydrolytic’ activity with a hyper-TG activity on DP2 to DP6 substrates, thus generating long chain COS of DP up to 13 and a hydrolytic property that breaks down DP8 and DP10 TG products to DP2 and DP4 COS respectively [ 24 ]. TG can however be of advantage if the chitinase has a hyper-TG activity like Enterobacter cloacae chitinase designated Ec Chi1, reported to produce long chain COS up to DP9 [ 84 ].…”

Section: Classification Based On Differences In Catalytic Mechanismsmentioning

confidence: 99%

Chitinase: diversity, limitations, and trends in engineering for suitable applications

2018

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Chitinases catalyze the degradation of chitin, a ubiquitous polymer generated from the cell walls of fungi, shells of crustaceans, and cuticles of insects. They are gaining increasing attention in medicine, agriculture, food and drug industries, and environmental management. Their roles in the degradation of chitin for the production of industrially useful products and in the control of fungal pathogens and insect pests render them attractive for such purposes. However, chitinases have diverse sources, characteristics, and mechanisms of action that seem to restrain optimization procedures and render standardization techniques for enhanced practical applications complex. Hence, results of laboratory trials are not usually consistent with real-life applications. With the growing field of protein engineering, these complexities can be overcome by modifying or redesigning chitinases to enhance specific features required for specific applications. In this review, the variations in features and mechanisms of chitinases that limit their exploitation in biotechnological applications are compiled. Recent attempts to engineer chitinases for improved efficiency are also highlighted.

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“…They unravelled how a 13-residue protruding loop in the deep active binding cleft blocked a large number of subsites and suggested this structural characteristic to be responsible for the TG aptitude [67]. Furthermore, the majority of dual action mode chitinases seem to exhibit aromatic amino residues in their catalytic site, suggesting a processive mechanism [67,68]. The exact mechanism of TG events is not fully understood, however for the catalysis of disaccharides it could be shown that the TG is autocondensation-driven and has to happen faster than the glycosidic hydrolysis [69].…”

Section: Chitinases With Transglycosylation Capabilitiesmentioning

confidence: 99%

“…Consistent evidence from several studies illustrates how the transfer of COS is dependent on the amount and length of substrate and the proportion and nature of the enzyme. Hereby an excess amount of substrate appears to be beneficial for a TG activity, while a surplus of the glycosidase seems to push the reaction equilibrium towards a predominant hydrolysis agitation [66,68,70].…”

Section: Chitinases With Transglycosylation Capabilitiesmentioning

confidence: 99%

Enzymatic Modification of Native Chitin and Conversion to Specialty Chemical Products

Arnold

Brück²,

Garbe

et al. 2020

Marine Drugs

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Chitin is one of the most abundant biomolecules on earth, occurring in crustacean shells and cell walls of fungi. While the polysaccharide is threatening to pollute coastal ecosystems in the form of accumulating shell-waste, it has the potential to be converted into highly profitable derivatives with applications in medicine, biotechnology, and wastewater treatment, among others. Traditionally this is still mostly done by the employment of aggressive chemicals, yielding low quality while producing toxic by-products. In the last decades, the enzymatic conversion of chitin has been on the rise, albeit still not on the same level of cost-effectiveness compared to the traditional methods due to its multi-step character. Another severe drawback of the biotechnological approach is the highly ordered structure of chitin, which renders it nigh impossible for most glycosidic hydrolases to act upon. So far, only the Auxiliary Activity 10 family (AA10), including lytic polysaccharide monooxygenases (LPMOs), is known to hydrolyse native recalcitrant chitin, which spares the expensive first step of chemical or mechanical pre-treatment to enlarge the substrate surface. The main advantages of enzymatic conversion of chitin over conventional chemical methods are the biocompability and, more strikingly, the higher product specificity, product quality, and yield of the process. Products with a higher Mw due to no unspecific depolymerisation besides an exactly defined degree and pattern of acetylation can be yielded. This provides a new toolset of thousands of new chitin and chitosan derivatives, as the physio-chemical properties can be modified according to the desired application. This review aims to provide an overview of the biotechnological tools currently at hand, as well as challenges and crucial steps to achieve the long-term goal of enzymatic conversion of native chitin into specialty chemical products.

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Transglycosylation by a chitinase from Enterobacter cloacae subsp. cloacae generates longer chitin oligosaccharides

Cited by 39 publications

References 61 publications

Molecular characterization of a novel chitinase CmChi1 from Chitinolyticbacter meiyuanensis SYBC-H1 and its use in N-acetyl-d-glucosamine production

Molecular characterization of a novel chitinase CmChi1 from Chitinolyticbacter meiyuanensis SYBC-H1 and its use in N-acetyl-d-glucosamine production

Chitinase: diversity, limitations, and trends in engineering for suitable applications

Enzymatic Modification of Native Chitin and Conversion to Specialty Chemical Products

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