Mammalian chitinases and the related chilectins （ChiLs） belong to the GH18 family, which hydrolyse the glycosidic bond of chitin by a substrate-assisted mechanism. Chitin the fundamental component in the coating of ...Mammalian chitinases and the related chilectins （ChiLs） belong to the GH18 family, which hydrolyse the glycosidic bond of chitin by a substrate-assisted mechanism. Chitin the fundamental component in the coating of numerous living species is the most abundant natural biopolymer. Mounting evidence suggest that the function of the majority of the mammalian chitinases is not exclusive to catalyze the hydrolysis of chitin producing pathogens, but include crucial role specifc in the immunologic activities. The chitinases and chitinase-like proteins are expressed in response to different proinflammatory cues in various tissues by activated macrophages, neutrophils and in different monocyte-derived cell lines. The mechanism and molecular interaction of chitinases in relation to immune regulation embrace bacterial infection, infammation, dismetabolic and degenerative disease. The aim of this review is to update the reader with regard to the role of chitinases proposed in the recent innate and adaptive immunity literature. The deep scrutiny of this family of enzymes could be a useful base for further studies addressed to the development of potential procedure directing these molecules as diagnostic and prognostic markers for numerous immune and infammatory diseases.展开更多
Chitinases play an important role in regulating plant growth and development, especially defending themselves from fungal pathogens. It is important to do the biological analyses in crops. In this study, the result sh...Chitinases play an important role in regulating plant growth and development, especially defending themselves from fungal pathogens. It is important to do the biological analyses in crops. In this study, the result showed that the chitinases were distributed into the whole genome in rice, and nearly the whole genome in maize expect for Chromosome 9. The clustering results showed that one out of three chitinases from maize and rice belonged to new groups, which were separated from those in the conformed Classes I-VII. The identification of most amino acid sequences was very low among the chitinases from rice and/or maize. It was inferred that the chitinases with different functions were relatively stable during plant evolution. The relationship of chitinases expression between leaf blade and anther was positively significant in maize, but not significant in rice. Additionally, the ratio of chitinases with up- or down-regulated expression in sensitive maize under Fusarium moniliforme inoculation was different from that in sensitive rice under Magnaporthe grisea inoculation. It might be result from different tissues infected by different fungi. The number of chitinases from resistant maize was less than that from sensitive maize, which inferred that the resistant pathways on F. moniliforme should be not only chitin induced Pathogen-associated molecular PTI （patterns-triggered immunity） pathway, but also might include other PTI pathways that improve tolerance to F. moniliforme. The analysis of expression pattern of chitinases from maize and rice under fungi inoculation will be contributed into further research on the defense mechanism of fungi in crops.展开更多
Chitinases （EC3.2.1.14）, found in a wide range of organisms, catalyze the hydrolysis of chitin and play a major role in defense mechanisms against fungal pathogens. The alignment and typical domains were analyzed us...Chitinases （EC3.2.1.14）, found in a wide range of organisms, catalyze the hydrolysis of chitin and play a major role in defense mechanisms against fungal pathogens. The alignment and typical domains were analyzed using basic local alignment search tool （BLAST） and simple modular architecture research tool （SMART）, respectively. On the basis of the annotations of flee （Oryza sativa L.） and Arabidopsis genomic sequences and using the bio-software SignalP3.0, TMHMM2.0, TargetPl.1, and big-Pi Predictor, 25 out of 37 and 16 out of 24 open reading frames （ORFs） with chitinase activity from rice and Arabidopsis, respectively, were predicted to have signal pepfides （SPs）, which have an average of 24.8 amino acids at the N-terminal region. Some of the chitinases were secreted extracellularly, whereas some were located in the vacuole. The phylogenic relationship was analyzed with 61 ORFs and 25 known ehitinases and they were classified into 6 clusters using Clustal X and MEGA3.1. This classification is not completely consistent when compared with the traditional system that classifies the chitinases into 7 classes. The frequency of distribution of amino acid residues was distinct in different clusters. The contents of alanine, glycine, serine, and leucine were very high in each cluster, whereas the contents of methionine, histidine, tryptophan, and cysteine were lower than 20%. Each cluster had distinct amino acid characteristics. Alanine, valine, leucine, cysteine, serine, and lysine were rich in Clusters Ⅰ to Ⅵ, respectively.展开更多
Chitinases catalyze the hydrolysis of chitin, a linear homopolymer of β-(1,4)-linked N-acetylglucosamine. The broad range of applications of chitinolytic enzymes makes their identification and study very promising. M...Chitinases catalyze the hydrolysis of chitin, a linear homopolymer of β-(1,4)-linked N-acetylglucosamine. The broad range of applications of chitinolytic enzymes makes their identification and study very promising. Metagenomic approaches offer access to functional genes in uncultured representatives of the microbiota and hold great potential in the discovery of novel enzymes, but tools to extensively explore these data are still scarce. In this study, we develop a chitinase mining pipeline to facilitate the comprehensive search of these enzymes in environmental metagenomic databases and also to explore phylogenetic relationships among the retrieved sequences. In order to perform the analyses, UniprotKB fungal and bacterial chitinases sequences belonging to the glycoside hydrolases (GH) family-18, 19 and 20 were used to generate 15 reference datasets, which were then used to generate high quality seed alignments with the MAFFT program. Profile Hidden Markov Models (pHMMs) were built from each seed alignment using the hmmbuild program of HMMER v3.0 package. The best-hit sequences returned by hmmsearch against two environmental metagenomic databases (Community Cyberinfrastructure for Advanced Microbial Ecology Research and Analysis—CAMERA and Integrated Microbial Genomes—IMG/M) were retrieved and further analyzed. The NJ trees generated for each chitinase dataset showed some variability in the catalytic domain region of the metagenomic sequences and revealed common sequence patterns among all the trees. The scanning of the retrieved metagenomic sequences for chitinase conserved domains/signatures using both the InterPro and the RPS-BLAST tools confirmed the efficacy and sensitivity of our pHMM-based approach in detecting putative chitinases sequences. These analyses provide insight into the potential reservoir of novel molecules in metagenomic databases while supporting the chitinase mining pipeline developed in this work. By using our chitinase mining pipeline, a larger number of previously unannotated metagenomic chitinase sequences can be classified, enabling further studies on these enzymes.展开更多
Vibrio pacini synthesizes multiple chitinases, of which three have been purified in this study by ammonium sulphate fractionation, chitin affinity chromatography and gel chromatography. Molecular weights of the three ...Vibrio pacini synthesizes multiple chitinases, of which three have been purified in this study by ammonium sulphate fractionation, chitin affinity chromatography and gel chromatography. Molecular weights of the three chitinases, Chi1, Chi2 and Chi3 are 27×103, 39×103 and 46×103 respectively, as determined by sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE). The purified enzymes have optimal activity at pH 7–8, and retain 50% enzymatic activity pH 4–9. The activities of chitinases are inhibited by Pb2+, Fe3+ and Cu2+, and increased by Ca2+, Mg2+ and Mn2+. Chi3 is found to inhibit the growth of six species of fungi. Such characters of chitinase are different from those of any other chitinase that were reported before. Key words chitinase - Vibrio pacini - purification - inhibit CLC number Q 55 Foundation item: Supported by the Key Technologies Research and Development Programme of the Tenth Five-Year Plan of the Nation Scientific and Technological Development (2001 BA708B04-07)Biography: HAN Bao-qin (1963-), female, Professor.展开更多
Application of crab shell chitin or pentamer chitin oligosaccharide to Arabidopsis seedlings increased toler- ance to salinity in wild-type but not in knockout mutants of the LysM Receptor-Like Kinasel （CERK1/LysM R...Application of crab shell chitin or pentamer chitin oligosaccharide to Arabidopsis seedlings increased toler- ance to salinity in wild-type but not in knockout mutants of the LysM Receptor-Like Kinasel （CERK1/LysM RLK1） gene, known to play a critical role in signaling defense responses induced by exogenous chitin. Arabidopsis plants overexpress- ing the endochitinase chit36 and hexoaminidase excyl genes from the fungus Trichoderma asperelleoides T203 showed increased tolerance to salinity, heavy-metal stresses, and Botrytis cinerea infection. Resistant lines, overexpressing fungal chitinases at different levels, were outcrossed to lysm rlkl mutants. Independent homozygous hybrids lost resistance to biotic and abiotic stresses, despite enhanced chitinase activity. Expression analysis of 270 stress-related genes, including those induced by reactive oxygen species （ROS） and chitin, revealed constant up-regulation （at least twofold） of 10 genes in the chitinase-overexpressing line and an additional 76 salt-induced genes whose expression was not elevated in the lysm rlkl knockout mutant or the hybrids harboring the mutation. These findings elucidate that chitin-induced signaling mediated by LysM RLK1 receptor is not limited to biotic stress response but also encompasses abiotic-stress signaling and can be conveyed by ectopic expression of chitinases in plants.展开更多
Chitin is an abundant aminopolysaccharide found in insect pests and phytopathogenic microorganisms but absent in higher plants and vertebrates. It is crucial for mitigating threats posed by chitin-containing organisms...Chitin is an abundant aminopolysaccharide found in insect pests and phytopathogenic microorganisms but absent in higher plants and vertebrates. It is crucial for mitigating threats posed by chitin-containing organisms to human health, food safety, and agriculture. Therefore, targeting the chitin biosynthesisassociated bioprocess holds a promise for developing human-safe and eco-friendly antifungal agents or pesticides. Chitin biosynthesis requires chitin synthase and associated factors, which are involved in the modification, regulation, organization or turnover of chitin during its biosynthesis. A number of enzymes such as chitinases, hexosaminidases, chitin deacetylases are closely related and therefore are promising targets for designing novel agrochemicals that target at chitin biosynthesis. This review summarizes the advances in understanding chitin biology over the past decade by our research group and collaborates,specifically regarding essential proteins linked to chitin biosynthesis that can be exploited as promising pesticide targets. Examples of small bioactive molecules that against the activity of these targets are given.展开更多
The biochemical response of Coffea arabica var.Borbon to chitosan and chitosan oligosaccharides(COS)was evaluated in one-year-old plants under greenhouse conditions.COS solutions were synthesized through chemical and ...The biochemical response of Coffea arabica var.Borbon to chitosan and chitosan oligosaccharides(COS)was evaluated in one-year-old plants under greenhouse conditions.COS solutions were synthesized through chemical and physical hydrolysis using acetic acid,hydrogen peroxide,and microwave irradiation.The obtained COS had an average molecular weight(Mw)of 3549.90±0.33 Daltons(Da),a deacetylation degree(DD)of 76.64±1.12%,and a polymerization degree(PD)of 18.91±0.0018.Solutions of chitosan and COS were applied to C.arabica var.Borbon at concentrations of 0.25,0.5,and 1 wt%.The experimental design was conducted using a completely randomized design with four replications.The biochemical responses assessed included soluble protein content,phenylalanine ammonia-lyase(PAL),chitinase,β-1,3-glucanase,peroxidase,catalase,and chlorophyll fluorescence.The application of COS demonstrated significant differences(α=0.05)in protein concentration,with the activity ofβ-1,3-glucanase,chitinase,and catalase being 1.5,7.5,and 3.9 times higher,respectively,while showing similar behavior to chitosan in PAL activity,both up to 4.4 times higher than the distilled water control and lower than chitosan in peroxidase activity.Treatments with chitosan yielded a higher photochemical efficiency of Photosystem II(PSII).The application of COS suggests a viable foliar alternative to active plant defense mechanisms without the risk of phytotoxicity.展开更多
Chitin is the second most abundant renewable polysaccharide on Earth.The degradation of chitin into soluble and bioactive N-acetyl chitooligosaccharides(NCOSs)and N-acetyl-D-glucosamine(GlcNAc)has emerged as a pivotal...Chitin is the second most abundant renewable polysaccharide on Earth.The degradation of chitin into soluble and bioactive N-acetyl chitooligosaccharides(NCOSs)and N-acetyl-D-glucosamine(GlcNAc)has emerged as a pivotal step in the efficient and sustainable utilization of chitin resources.However,because of its dense structure,high crystallinity,and poor solubility,chitin typically needs pretreatment via chemical,physical,and other methods before enzymatic conversion to enhance the accessibility between substrates and enzyme molecules.Consequently,there has been considerable interest in exploring the direct biological degradation of crystalline chitin as a cost-effective and environment-friendly technology.This review endeavors to present several biological methods for the direct degradation of chitin.We primarily focused on the importance of chitinase containing chitin-binding domain(CBD).Additionally,various modification strategies for increasing the degradation efficiency of crystalline chitin were introduced.Subsequently,the review systematically elucidated critical components of multi-enzyme catalytic systems,highlighting their potential for chitin degradation.Furthermore,the application of microorganisms in the degradation of crystalline chitin was also discussed.The insights in this review contribute to the explorations and investigations of enzymatic and microbial approaches for the direct degradation of crystalline chitin,thereby fostering advancements in biomass conversion.展开更多
Thermophilic endo-chitinases are essential for production of highly polymerized chitooligosaccharides,which are advantageous for plant immunity,animal nutrition and health.However,thermophilic endo-chitinases are scar...Thermophilic endo-chitinases are essential for production of highly polymerized chitooligosaccharides,which are advantageous for plant immunity,animal nutrition and health.However,thermophilic endo-chitinases are scarce and the transformation from exo-to endo-activity of chitinases is still a challenging problem.In this study,to enhance the endo-activity of the thermophilic chitinase Chi304,we proposed two approaches for rational design based on comprehensive structural and evolutionary analyses.Four effective single-point mutants were identified among 28 designed mutations.The ratio of(GlcNAc)3 to(GlcNAc)2 quantity(DP3/2)in the hydrolysates of the four single-point mutants undertaking colloidal chitin degradation were 1.89,1.65,1.24,and 1.38 times that of Chi304,respectively.When combining to double-point mutants,the DP3/2 proportions produced by F79A/W140R,F79A/M264L,F79A/W272R,and M264L/W272R were 2.06,1.67,1.82,and 1.86 times that of Chi304 and all four double-point mutants exhibited enhanced endo-activity.When applied to produce chitooligosaccharides(DP≥3),F79A/W140R accumulated the most(GlcNAc)4,while M264L/W272R was the best to produce(GlcNAc)3,which was 2.28 times that of Chi304.The two mutants had exposed shallower substrate-binding pockets and stronger binding abilities to shape the substrate.Overall,this research offers a practical approach to altering the cutting pattern of a chitinase to generate functional chitooligosaccharides.展开更多
[Objective] This study aimed to screen endophytic bacteria which is antag- onistic to cotton Fusarium wilt. [Method] Fresh cotton plants collected from cotton- growing areas in Jingzhou City, Hubei Province were used ...[Objective] This study aimed to screen endophytic bacteria which is antag- onistic to cotton Fusarium wilt. [Method] Fresh cotton plants collected from cotton- growing areas in Jingzhou City, Hubei Province were used as experimental materials to isolate endophytic bacteria. Through chitinase test and co-culturing both micro-or- ganisms side by side on the same PDA culture plate, antagonistic strains to cotton Fusarium wilt were screened. [Result] A total of 83 bacterial isolates were obtained from cotton plants grown in the fields, six of which were chitinase-productive bacte- ria. Through chitinase test and co-culturing both micro-organisms side by side on the same PDA culture plate, strain V-8 which had the strongest antagonistic effect on Fusarium oxysporum f. sp. vasinfectum was screened. Strain V-8 had a wider anti- fungal spectrum with certain inhibitory effect on all the six important pathogenic fungi including Fusarium oxysporum f. sp niveum; it colonized stably in the rhizospheric soil of cotton, with a colonization density of up to 6.2x10s cfu/g fifty days after inoc- ulation; the relative effect on controlling cotton Fusarium wilt in pot test was 73.2%. The Findings of this study suggested that strain V-8 had great potential for biological control of cotton Fusarium wilt and could be taken as a substantial material for the cloning of chitinase genes. [Conclusion] The results from this study provides bases for the control of cotton fusarium wilt, as well as the exploitation of endophytic bac- teria resources in cotton and the development of novel biological pesticides.展开更多
Tebufenozide,an efficient insect growth regulator(IGR)against lepidopteran pests,presents a novel mode of action with minimal non-target impact.By competing with ecdysteroids for ecdysone receptor(EcR)binding,it regul...Tebufenozide,an efficient insect growth regulator(IGR)against lepidopteran pests,presents a novel mode of action with minimal non-target impact.By competing with ecdysteroids for ecdysone receptor(EcR)binding,it regulates insect growth precisely.This study explores tebufenozide's potential as a multitarget IGR,targeting both EcR and Ostrinia furnacalis chitinase I(OfChtI).The inhibitory activity against OfChtI is comparable to that of substrates(GlcNAc)5,with an IC50 of 45.77μM.Our computational findings indicate that tebufenozide binds at the subsite1 to t1 of OfChtI through various interactions.Notably,tebufenozide establishes a pi-pi interaction with the flipped sidechain of Trp107,enabling tebufenozide to deeply penetrate into the S1 pocket,thereby obstructing substrate binding to OfChtI.These insights highlight the potency of multitarget strategies,laying the groundwork for innovative IGR designs that offer comprehensive pest management solutions.展开更多
The different resistance of cotton (Gossypium hirsutum L.) cultivars to crude toxin of Verticillium dah/iae(VD) was correlated with the activities of chitinase and β-1, 3-glucanase in callus cells. The activities of ...The different resistance of cotton (Gossypium hirsutum L.) cultivars to crude toxin of Verticillium dah/iae(VD) was correlated with the activities of chitinase and β-1, 3-glucanase in callus cells. The activities of chitinase and β-1, 3-glucanase in the callus cells treated with the VD-toxin were increased to the higher level at earlier time point in resistant cultivars than these in the susceptible cultivars. Exogenous salicylic acid (SA) induced the accumulation of chitinase and β -1,3-glucanase, which resulted in the resistance of callus cells to the VD. toxin. Western blot using a polyclonal antibody against β -1,3-glucanase identified 28 kD protein that was induced by VD-toxin, SA, or VD-toxin plus SA.展开更多
[Objective] The aim of this study was to isolate chitinase gene from Trichoderma atroviride strain SS003. [Method] With the aeciospore wall of armandii pine blister rust as inducer, chitinase gene was induced to expre...[Objective] The aim of this study was to isolate chitinase gene from Trichoderma atroviride strain SS003. [Method] With the aeciospore wall of armandii pine blister rust as inducer, chitinase gene was induced to express in Trichoderma atroviride cells. The cDNA fragment of chitinase gene was cloned by RT-PCR approach. [Result] The activity of chitinase induced reached 40.17 μg/10 min; and the specific fragment amplified was 834 bp in length and proved to be the fragment of chitinase gene by sequencing and sequence analysis. [Conclusion] The result showed the feasibility of isolating the full length of chitinase gene and its transformation, and further producing chitinase.展开更多
[ Objective] The aim of this study was to clone Class Ⅱ chitinase gene in Leymus chinensis grown in saline land in Heilongjiang Province and analyze its sequence, which provided a foundation for further study on the ...[ Objective] The aim of this study was to clone Class Ⅱ chitinase gene in Leymus chinensis grown in saline land in Heilongjiang Province and analyze its sequence, which provided a foundation for further study on the biological function and application of chitinasa gene. [ Method] cDNA library of Leymus chinensis leaves were constructed, and its DNA sequence was determined or analyzed, while the homology of chitinasa gene and amino acid sequence was compared with that in GenBank. [ Result] One full length cDNA fragment with length of 996 bp was cloned from cDNA library of Leymus chinensis leaves. The length of ORF was 768 bp encoding 225 amino acids （GenBank accession number： EU344908）. The encoding products lacked CBD and C-terminal extension region from the view of structure, but had structural characters of Class Ⅱ chitinase gene, which indicated that amino acid sequence had high homology compared with Class Ⅱ chitinase gene of rye and wheat. The constructed recombinant vector pQE-LcChi2 could express a protein of 27 kD through induction, which was consistent with the deduced encoding product of pQE-LcChi2 gene. [ Conclusion] LcChl2 gene is an expression gene, which can express in E. coll.展开更多
DA novel chitinase gene (GhChia7) was isolated from salicylic acid (SA)-treated cotton cotyledons and characterized by DNA sequence analysis of its cDNA and genomic DNA clone. The deduced amino acid sequence, designat...DA novel chitinase gene (GhChia7) was isolated from salicylic acid (SA)-treated cotton cotyledons and characterized by DNA sequence analysis of its cDNA and genomic DNA clone. The deduced amino acid sequence, designated as class VII chitinase, shares about 30% identity to class I or II chitinases, and does not correspond to any of the previously characterized classes I-VI chitinases. Northern blotting analysis showed that the transcripts of GhChia7 were abundant both in cotton fibers and in the roots of the seedlings. The accumulation of GhChia7 mRNA in SA-treated cotyledons reached maximum at 7.5 mmol/ L concentration after 18 h. Results indicate that GhChia7 might play an important role in cotton's active defense response.展开更多
文摘Mammalian chitinases and the related chilectins （ChiLs） belong to the GH18 family, which hydrolyse the glycosidic bond of chitin by a substrate-assisted mechanism. Chitin the fundamental component in the coating of numerous living species is the most abundant natural biopolymer. Mounting evidence suggest that the function of the majority of the mammalian chitinases is not exclusive to catalyze the hydrolysis of chitin producing pathogens, but include crucial role specifc in the immunologic activities. The chitinases and chitinase-like proteins are expressed in response to different proinflammatory cues in various tissues by activated macrophages, neutrophils and in different monocyte-derived cell lines. The mechanism and molecular interaction of chitinases in relation to immune regulation embrace bacterial infection, infammation, dismetabolic and degenerative disease. The aim of this review is to update the reader with regard to the role of chitinases proposed in the recent innate and adaptive immunity literature. The deep scrutiny of this family of enzymes could be a useful base for further studies addressed to the development of potential procedure directing these molecules as diagnostic and prognostic markers for numerous immune and infammatory diseases.
基金Acknowledgments This work was supported by Specialized Research Fund for New Teachers of Doctoral Program in the University of China （No. 20125103120011） and the Natural National Science Foundation of China （No. 31201274）.
文摘Chitinases play an important role in regulating plant growth and development, especially defending themselves from fungal pathogens. It is important to do the biological analyses in crops. In this study, the result showed that the chitinases were distributed into the whole genome in rice, and nearly the whole genome in maize expect for Chromosome 9. The clustering results showed that one out of three chitinases from maize and rice belonged to new groups, which were separated from those in the conformed Classes I-VII. The identification of most amino acid sequences was very low among the chitinases from rice and/or maize. It was inferred that the chitinases with different functions were relatively stable during plant evolution. The relationship of chitinases expression between leaf blade and anther was positively significant in maize, but not significant in rice. Additionally, the ratio of chitinases with up- or down-regulated expression in sensitive maize under Fusarium moniliforme inoculation was different from that in sensitive rice under Magnaporthe grisea inoculation. It might be result from different tissues infected by different fungi. The number of chitinases from resistant maize was less than that from sensitive maize, which inferred that the resistant pathways on F. moniliforme should be not only chitin induced Pathogen-associated molecular PTI （patterns-triggered immunity） pathway, but also might include other PTI pathways that improve tolerance to F. moniliforme. The analysis of expression pattern of chitinases from maize and rice under fungi inoculation will be contributed into further research on the defense mechanism of fungi in crops.
基金This work was supported by the 863 Program (No. 2002AA245041), the National Natural Science Foundation of China (No. 30260006), and the R&D Foundation of Yunnan Province (No. 2003GP06).
文摘Chitinases （EC3.2.1.14）, found in a wide range of organisms, catalyze the hydrolysis of chitin and play a major role in defense mechanisms against fungal pathogens. The alignment and typical domains were analyzed using basic local alignment search tool （BLAST） and simple modular architecture research tool （SMART）, respectively. On the basis of the annotations of flee （Oryza sativa L.） and Arabidopsis genomic sequences and using the bio-software SignalP3.0, TMHMM2.0, TargetPl.1, and big-Pi Predictor, 25 out of 37 and 16 out of 24 open reading frames （ORFs） with chitinase activity from rice and Arabidopsis, respectively, were predicted to have signal pepfides （SPs）, which have an average of 24.8 amino acids at the N-terminal region. Some of the chitinases were secreted extracellularly, whereas some were located in the vacuole. The phylogenic relationship was analyzed with 61 ORFs and 25 known ehitinases and they were classified into 6 clusters using Clustal X and MEGA3.1. This classification is not completely consistent when compared with the traditional system that classifies the chitinases into 7 classes. The frequency of distribution of amino acid residues was distinct in different clusters. The contents of alanine, glycine, serine, and leucine were very high in each cluster, whereas the contents of methionine, histidine, tryptophan, and cysteine were lower than 20%. Each cluster had distinct amino acid characteristics. Alanine, valine, leucine, cysteine, serine, and lysine were rich in Clusters Ⅰ to Ⅵ, respectively.
文摘Chitinases catalyze the hydrolysis of chitin, a linear homopolymer of β-(1,4)-linked N-acetylglucosamine. The broad range of applications of chitinolytic enzymes makes their identification and study very promising. Metagenomic approaches offer access to functional genes in uncultured representatives of the microbiota and hold great potential in the discovery of novel enzymes, but tools to extensively explore these data are still scarce. In this study, we develop a chitinase mining pipeline to facilitate the comprehensive search of these enzymes in environmental metagenomic databases and also to explore phylogenetic relationships among the retrieved sequences. In order to perform the analyses, UniprotKB fungal and bacterial chitinases sequences belonging to the glycoside hydrolases (GH) family-18, 19 and 20 were used to generate 15 reference datasets, which were then used to generate high quality seed alignments with the MAFFT program. Profile Hidden Markov Models (pHMMs) were built from each seed alignment using the hmmbuild program of HMMER v3.0 package. The best-hit sequences returned by hmmsearch against two environmental metagenomic databases (Community Cyberinfrastructure for Advanced Microbial Ecology Research and Analysis—CAMERA and Integrated Microbial Genomes—IMG/M) were retrieved and further analyzed. The NJ trees generated for each chitinase dataset showed some variability in the catalytic domain region of the metagenomic sequences and revealed common sequence patterns among all the trees. The scanning of the retrieved metagenomic sequences for chitinase conserved domains/signatures using both the InterPro and the RPS-BLAST tools confirmed the efficacy and sensitivity of our pHMM-based approach in detecting putative chitinases sequences. These analyses provide insight into the potential reservoir of novel molecules in metagenomic databases while supporting the chitinase mining pipeline developed in this work. By using our chitinase mining pipeline, a larger number of previously unannotated metagenomic chitinase sequences can be classified, enabling further studies on these enzymes.
文摘Vibrio pacini synthesizes multiple chitinases, of which three have been purified in this study by ammonium sulphate fractionation, chitin affinity chromatography and gel chromatography. Molecular weights of the three chitinases, Chi1, Chi2 and Chi3 are 27×103, 39×103 and 46×103 respectively, as determined by sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE). The purified enzymes have optimal activity at pH 7–8, and retain 50% enzymatic activity pH 4–9. The activities of chitinases are inhibited by Pb2+, Fe3+ and Cu2+, and increased by Ca2+, Mg2+ and Mn2+. Chi3 is found to inhibit the growth of six species of fungi. Such characters of chitinase are different from those of any other chitinase that were reported before. Key words chitinase - Vibrio pacini - purification - inhibit CLC number Q 55 Foundation item: Supported by the Key Technologies Research and Development Programme of the Tenth Five-Year Plan of the Nation Scientific and Technological Development (2001 BA708B04-07)Biography: HAN Bao-qin (1963-), female, Professor.
文摘Application of crab shell chitin or pentamer chitin oligosaccharide to Arabidopsis seedlings increased toler- ance to salinity in wild-type but not in knockout mutants of the LysM Receptor-Like Kinasel （CERK1/LysM RLK1） gene, known to play a critical role in signaling defense responses induced by exogenous chitin. Arabidopsis plants overexpress- ing the endochitinase chit36 and hexoaminidase excyl genes from the fungus Trichoderma asperelleoides T203 showed increased tolerance to salinity, heavy-metal stresses, and Botrytis cinerea infection. Resistant lines, overexpressing fungal chitinases at different levels, were outcrossed to lysm rlkl mutants. Independent homozygous hybrids lost resistance to biotic and abiotic stresses, despite enhanced chitinase activity. Expression analysis of 270 stress-related genes, including those induced by reactive oxygen species （ROS） and chitin, revealed constant up-regulation （at least twofold） of 10 genes in the chitinase-overexpressing line and an additional 76 salt-induced genes whose expression was not elevated in the lysm rlkl knockout mutant or the hybrids harboring the mutation. These findings elucidate that chitin-induced signaling mediated by LysM RLK1 receptor is not limited to biotic stress response but also encompasses abiotic-stress signaling and can be conveyed by ectopic expression of chitinases in plants.
基金supported by the National Key Research and Development Program of China (No. 2022YFD1700200)the National Natural Science Foundation of China (Nos. 32161133010, 3230170969)+1 种基金the Innovation Program of Chinese Academy of Agricultural Sciences, the Shenzhen Science and Technology Program (No. KQTD20180411143628272)the Special Funds for Science Technology Innovation and Industrial Development of Shenzhen Dapeng New District (No. PT202101–02)。
文摘Chitin is an abundant aminopolysaccharide found in insect pests and phytopathogenic microorganisms but absent in higher plants and vertebrates. It is crucial for mitigating threats posed by chitin-containing organisms to human health, food safety, and agriculture. Therefore, targeting the chitin biosynthesisassociated bioprocess holds a promise for developing human-safe and eco-friendly antifungal agents or pesticides. Chitin biosynthesis requires chitin synthase and associated factors, which are involved in the modification, regulation, organization or turnover of chitin during its biosynthesis. A number of enzymes such as chitinases, hexosaminidases, chitin deacetylases are closely related and therefore are promising targets for designing novel agrochemicals that target at chitin biosynthesis. This review summarizes the advances in understanding chitin biology over the past decade by our research group and collaborates,specifically regarding essential proteins linked to chitin biosynthesis that can be exploited as promising pesticide targets. Examples of small bioactive molecules that against the activity of these targets are given.
文摘The biochemical response of Coffea arabica var.Borbon to chitosan and chitosan oligosaccharides(COS)was evaluated in one-year-old plants under greenhouse conditions.COS solutions were synthesized through chemical and physical hydrolysis using acetic acid,hydrogen peroxide,and microwave irradiation.The obtained COS had an average molecular weight(Mw)of 3549.90±0.33 Daltons(Da),a deacetylation degree(DD)of 76.64±1.12%,and a polymerization degree(PD)of 18.91±0.0018.Solutions of chitosan and COS were applied to C.arabica var.Borbon at concentrations of 0.25,0.5,and 1 wt%.The experimental design was conducted using a completely randomized design with four replications.The biochemical responses assessed included soluble protein content,phenylalanine ammonia-lyase(PAL),chitinase,β-1,3-glucanase,peroxidase,catalase,and chlorophyll fluorescence.The application of COS demonstrated significant differences(α=0.05)in protein concentration,with the activity ofβ-1,3-glucanase,chitinase,and catalase being 1.5,7.5,and 3.9 times higher,respectively,while showing similar behavior to chitosan in PAL activity,both up to 4.4 times higher than the distilled water control and lower than chitosan in peroxidase activity.Treatments with chitosan yielded a higher photochemical efficiency of Photosystem II(PSII).The application of COS suggests a viable foliar alternative to active plant defense mechanisms without the risk of phytotoxicity.
基金supported by the National Key Research and Development Program of China(No.2023YFD2401504)the National Natural Science Foundation of China(Nos.U21A20271,32225039)+2 种基金the Key R&D Program of Shandong Province(No.2022TZXD001)the Earmarked Fund for CARS(No.CARS-48)the Qingdao Shinan District Science and Technology Plan Project(No.2022-3-010-SW).
文摘Chitin is the second most abundant renewable polysaccharide on Earth.The degradation of chitin into soluble and bioactive N-acetyl chitooligosaccharides(NCOSs)and N-acetyl-D-glucosamine(GlcNAc)has emerged as a pivotal step in the efficient and sustainable utilization of chitin resources.However,because of its dense structure,high crystallinity,and poor solubility,chitin typically needs pretreatment via chemical,physical,and other methods before enzymatic conversion to enhance the accessibility between substrates and enzyme molecules.Consequently,there has been considerable interest in exploring the direct biological degradation of crystalline chitin as a cost-effective and environment-friendly technology.This review endeavors to present several biological methods for the direct degradation of chitin.We primarily focused on the importance of chitinase containing chitin-binding domain(CBD).Additionally,various modification strategies for increasing the degradation efficiency of crystalline chitin were introduced.Subsequently,the review systematically elucidated critical components of multi-enzyme catalytic systems,highlighting their potential for chitin degradation.Furthermore,the application of microorganisms in the degradation of crystalline chitin was also discussed.The insights in this review contribute to the explorations and investigations of enzymatic and microbial approaches for the direct degradation of crystalline chitin,thereby fostering advancements in biomass conversion.
基金supported by the National Key R&D Program of China[Grant No.2021YFC2103002]the National Natural Science Foundation of China[Grant No.32202720]+1 种基金The Agricultural Science and Technology Innovation Program(CAAS-ZDRW202304)the China Agriculture Research System of MOF and MARA(CARS-41).
文摘Thermophilic endo-chitinases are essential for production of highly polymerized chitooligosaccharides,which are advantageous for plant immunity,animal nutrition and health.However,thermophilic endo-chitinases are scarce and the transformation from exo-to endo-activity of chitinases is still a challenging problem.In this study,to enhance the endo-activity of the thermophilic chitinase Chi304,we proposed two approaches for rational design based on comprehensive structural and evolutionary analyses.Four effective single-point mutants were identified among 28 designed mutations.The ratio of(GlcNAc)3 to(GlcNAc)2 quantity(DP3/2)in the hydrolysates of the four single-point mutants undertaking colloidal chitin degradation were 1.89,1.65,1.24,and 1.38 times that of Chi304,respectively.When combining to double-point mutants,the DP3/2 proportions produced by F79A/W140R,F79A/M264L,F79A/W272R,and M264L/W272R were 2.06,1.67,1.82,and 1.86 times that of Chi304 and all four double-point mutants exhibited enhanced endo-activity.When applied to produce chitooligosaccharides(DP≥3),F79A/W140R accumulated the most(GlcNAc)4,while M264L/W272R was the best to produce(GlcNAc)3,which was 2.28 times that of Chi304.The two mutants had exposed shallower substrate-binding pockets and stronger binding abilities to shape the substrate.Overall,this research offers a practical approach to altering the cutting pattern of a chitinase to generate functional chitooligosaccharides.
基金Supported by the Research Fund for Mid-career and Young Scientists of Education Department of Hubei Province(Q2011130)~~
文摘[Objective] This study aimed to screen endophytic bacteria which is antag- onistic to cotton Fusarium wilt. [Method] Fresh cotton plants collected from cotton- growing areas in Jingzhou City, Hubei Province were used as experimental materials to isolate endophytic bacteria. Through chitinase test and co-culturing both micro-or- ganisms side by side on the same PDA culture plate, antagonistic strains to cotton Fusarium wilt were screened. [Result] A total of 83 bacterial isolates were obtained from cotton plants grown in the fields, six of which were chitinase-productive bacte- ria. Through chitinase test and co-culturing both micro-organisms side by side on the same PDA culture plate, strain V-8 which had the strongest antagonistic effect on Fusarium oxysporum f. sp. vasinfectum was screened. Strain V-8 had a wider anti- fungal spectrum with certain inhibitory effect on all the six important pathogenic fungi including Fusarium oxysporum f. sp niveum; it colonized stably in the rhizospheric soil of cotton, with a colonization density of up to 6.2x10s cfu/g fifty days after inoc- ulation; the relative effect on controlling cotton Fusarium wilt in pot test was 73.2%. The Findings of this study suggested that strain V-8 had great potential for biological control of cotton Fusarium wilt and could be taken as a substantial material for the cloning of chitinase genes. [Conclusion] The results from this study provides bases for the control of cotton fusarium wilt, as well as the exploitation of endophytic bac- teria resources in cotton and the development of novel biological pesticides.
基金financially supported by the National Natural Science Foundation of China(No.22177132 and 21977114).
文摘Tebufenozide,an efficient insect growth regulator(IGR)against lepidopteran pests,presents a novel mode of action with minimal non-target impact.By competing with ecdysteroids for ecdysone receptor(EcR)binding,it regulates insect growth precisely.This study explores tebufenozide's potential as a multitarget IGR,targeting both EcR and Ostrinia furnacalis chitinase I(OfChtI).The inhibitory activity against OfChtI is comparable to that of substrates(GlcNAc)5,with an IC50 of 45.77μM.Our computational findings indicate that tebufenozide binds at the subsite1 to t1 of OfChtI through various interactions.Notably,tebufenozide establishes a pi-pi interaction with the flipped sidechain of Trp107,enabling tebufenozide to deeply penetrate into the S1 pocket,thereby obstructing substrate binding to OfChtI.These insights highlight the potency of multitarget strategies,laying the groundwork for innovative IGR designs that offer comprehensive pest management solutions.
文摘The different resistance of cotton (Gossypium hirsutum L.) cultivars to crude toxin of Verticillium dah/iae(VD) was correlated with the activities of chitinase and β-1, 3-glucanase in callus cells. The activities of chitinase and β-1, 3-glucanase in the callus cells treated with the VD-toxin were increased to the higher level at earlier time point in resistant cultivars than these in the susceptible cultivars. Exogenous salicylic acid (SA) induced the accumulation of chitinase and β -1,3-glucanase, which resulted in the resistance of callus cells to the VD. toxin. Western blot using a polyclonal antibody against β -1,3-glucanase identified 28 kD protein that was induced by VD-toxin, SA, or VD-toxin plus SA.
基金Supported by Science Foundation from Southwest Forestry College(200524M)Natural Science Foundation of Yunan Province(2002C0047M)Key Scientific and Technological Project of Yunan Province(2003NG12)~~
文摘[Objective] The aim of this study was to isolate chitinase gene from Trichoderma atroviride strain SS003. [Method] With the aeciospore wall of armandii pine blister rust as inducer, chitinase gene was induced to express in Trichoderma atroviride cells. The cDNA fragment of chitinase gene was cloned by RT-PCR approach. [Result] The activity of chitinase induced reached 40.17 μg/10 min; and the specific fragment amplified was 834 bp in length and proved to be the fragment of chitinase gene by sequencing and sequence analysis. [Conclusion] The result showed the feasibility of isolating the full length of chitinase gene and its transformation, and further producing chitinase.
基金Supported by Science and Technology Research Project of Education Department of Liaoning Province(2008120)IntroducedTalent Start-up Fund Project of Dalian Nationalities University(20056209)~~
文摘[ Objective] The aim of this study was to clone Class Ⅱ chitinase gene in Leymus chinensis grown in saline land in Heilongjiang Province and analyze its sequence, which provided a foundation for further study on the biological function and application of chitinasa gene. [ Method] cDNA library of Leymus chinensis leaves were constructed, and its DNA sequence was determined or analyzed, while the homology of chitinasa gene and amino acid sequence was compared with that in GenBank. [ Result] One full length cDNA fragment with length of 996 bp was cloned from cDNA library of Leymus chinensis leaves. The length of ORF was 768 bp encoding 225 amino acids （GenBank accession number： EU344908）. The encoding products lacked CBD and C-terminal extension region from the view of structure, but had structural characters of Class Ⅱ chitinase gene, which indicated that amino acid sequence had high homology compared with Class Ⅱ chitinase gene of rye and wheat. The constructed recombinant vector pQE-LcChi2 could express a protein of 27 kD through induction, which was consistent with the deduced encoding product of pQE-LcChi2 gene. [ Conclusion] LcChl2 gene is an expression gene, which can express in E. coll.
文摘DA novel chitinase gene (GhChia7) was isolated from salicylic acid (SA)-treated cotton cotyledons and characterized by DNA sequence analysis of its cDNA and genomic DNA clone. The deduced amino acid sequence, designated as class VII chitinase, shares about 30% identity to class I or II chitinases, and does not correspond to any of the previously characterized classes I-VI chitinases. Northern blotting analysis showed that the transcripts of GhChia7 were abundant both in cotton fibers and in the roots of the seedlings. The accumulation of GhChia7 mRNA in SA-treated cotyledons reached maximum at 7.5 mmol/ L concentration after 18 h. Results indicate that GhChia7 might play an important role in cotton's active defense response.
