Showing papers on "Chitin published in 2006"
TL;DR: There is strong evidence that the chitin, glucans and glycoproteins are covalently cross‐linked together and that the cross‐linking is a dynamic process that occurs extracellularly.
Abstract: The fungal cell wall is a dynamic structure that protects the cell from changes in osmotic pressure and other environmental stresses, while allowing the fungal cell to interact with its environment. The structure and biosynthesis of a fungal cell wall is unique to the fungi, and is therefore an excellent target for the development of anti-fungal drugs. The structure of the fungal cell wall and the drugs that target its biosynthesis are reviewed. Based on studies in a number of fungi, the cell wall has been shown to be primarily composed of chitin, glucans, mannans and glycoproteins. The biosynthesis of the various components of the fungal cell wall and the importance of the components in the formation of a functional cell wall, as revealed through mutational analyses, are discussed. There is strong evidence that the chitin, glucans and glycoproteins are covalently cross-linked together and that the cross-linking is a dynamic process that occurs extracellularly.
1,038 citations
TL;DR: This review deals with the fundamental aspects of chitin and chitosan such as the preparation of chItosan, crystallography, extent of N-acetylation, and some properties, and recent progress of their chemistry is discussed.
Abstract: Chitin and chitosan, typical marine polysaccharides as well as abundant biomass resources, are attracting a great deal of attention because of their distinctive biological and physicochemical characteristics. To fully explore the high potential of these specialty biopolymers, basic and application researches are being made extensively. This review deals with the fundamental aspects of chitin and chitosan such as the preparation of chitin and chitosan, crystallography, extent of N-acetylation, and some properties. Recent progress of their chemistry is then discussed, focusing on elemental modification reactions including acylation, alkylation, Schiff base formation and reductive alkylation, carboxyalkylation, phthaloylation, silylation, tosylation, quaternary salt formation, and sulfation and thiolation.
1,034 citations
TL;DR: As a nontoxic biodegradable material, as well as an elicitor, chitosan has the potential to become a new class of plant protectant, assisting towards the goal of sustainable agriculture.
687 citations
TL;DR: This review discusses the occurrence and structure of chitin, the types and sources of ch itinases, their mode of action, chit inase production, as well as molecular cloning and protein engineering of Chitinases and their biotechnological applications.
Abstract: Chitin and chitinases (EC 3.2.1.14) have an immense potential. Chitinolytic enzymes have wide-ranging applications such as preparation of pharmaceutically important chitooligosaccharides and N-acetyl D-glucosamine, preparation of single-cell protein, isolation of protoplasts from fungi and yeast, control of pathogenic fungi, treatment of chitinous waste, and control of malaria transmission. In this review, we discuss the occurrence and structure of chitin, the types and sources of chitinases, their mode of action, chitinase production, as well as molecular cloning and protein engineering of chitinases and their biotechnological applications.
499 citations
TL;DR: Recent advances in elucidating the structure, regulation and function of insect chitin synthases as they relate to what is known about fungal chit in synthases and other glycosyltransferases are summarized.
Abstract: Chitin is the most widespread amino polysaccharide in nature. The annual global amount of chitin is believed to be only one order of magnitude less than that of cellulose. It is a linear polymer composed of N-acetylglucosamines that are joined in a reaction catalyzed by the membrane-integral enzyme chitin synthase, a member of the family 2 of glycosyltransferases. The polymerization requires UDP–N-acetylglucosamines as a substrate and divalent cations as co-factors. Chitin formation can be divided into three distinct steps. In the first step, the enzymes‘ catalytic domain facing the cytoplasmic site forms the polymer. The second step involves the translocation of the nascent polymer across the membrane and its release into the extracellular space. The third step completes the process as single polymers spontaneously assemble to form crystalline microfibrils. In subsequent reactions the microfibrils combine with other sugars, proteins, glycoproteins and proteoglycans to form fungal septa and cell walls as well as arthropod cuticles and peritrophic matrices, notably in crustaceans and insects. In spite of the good effort by a hardy few, our present knowledge of the structure, topology and catalytic mechanism of chitin synthases is rather limited. Gaps remain in understanding chitin synthase biosynthesis, enzyme trafficking, regulation of enzyme activity, translocation of chitin chains across cell membranes, fibrillogenesis and the interaction of microfibrils with other components of the extracellular matrix. However, cumulating genomic data on chitin synthase genes and new experimental approaches allow increasingly clearer views of chitin synthase function and its regulation, and consequently chitin biosynthesis. In the present review, I will summarize recent advances in elucidating the structure, regulation and function of insect chitin synthases as they relate to what is known about fungal chitin synthases and other glycosyltransferases.
401 citations
TL;DR: In situ fluorescence studies showed that Avr4 also binds to cell walls of C. fulvum during infection of tomato, where it most likely protects the fungus against tomato chitinases, suggesting that AvR4 is a counter-defensive virulence factor.
Abstract: Resistance against the leaf mold fungus Cladosporium fulvum is mediated by the tomato Cf proteins which belong to the class of receptor-like proteins and indirectly recognize extracellular avirulence proteins (Avrs) of the fungus. Apart from triggering disease resistance, Avrs are believed to play a role in pathogenicity or virulence of C. fulvum. Here, we report on the avirulence protein Avr4, which is a chitin-binding lectin containing an invertebrate chitin-binding domain (CBM14). This domain is found in many eukaryotes, but has not yet been described in fungal or plant genomes. We found that interaction of Avr4 with chitin is specific, because it does not interact with other cell wall polysaccharides. Avr4 binds to chitin oligomers with a minimal length of three N-acetyl glucosamine residues. In vitro, Avr4 protects chitin against hydrolysis by plant chitinases. Avr4 also binds to chitin in cell walls of the fungi Trichoderma viride and Fusarium solani f. sp. phaseoli and protects these fungi against normally deleterious concentrations of plant chitinases. In situ fluorescence studies showed that Avr4 also binds to cell walls of C. fulvum during infection of tomato, where it most likely protects the fungus against tomato chitinases, suggesting that Avr4 is a counter-defensive virulence factor.
392 citations
TL;DR: In this article, a novel dissolving process for chitin and chitosan was developed by using the ionic liquid 1-butyl-3-methyl-imidazolium chloride ([Bmim]Cl) as a solvent.
389 citations
TL;DR: Chitin and chitosan were extracted from silkworm chrysalides and used for the production of a high purity and porous chiton, as observed by scanning electron microscopy (SEM) as discussed by the authors.
379 citations
TL;DR: Dietary intake of chitosan enhances the innate immune system and survivability of common carp in ponds and the role of ch itosan, chitin and levamisole as immunostimulants with regard to protection against A. hydrophila infection is discussed.
332 citations
TL;DR: Chitin and the matrix proteins Serp and Verm limit tube elongation, and it suggests that tube length is controlled independently of diameter by modulating physical properties of the chitin ECM, presumably by N-deacetylation of chit in and conversion to chitosan.
301 citations
TL;DR: The results indicate that the Chi-N, alone or with extracellular matrix proteins (particularly type I collagen), could be potential candidates for the cell attachment and spreading of normal human keratinocytes and fibroblasts.
TL;DR: The chitin hydrogel was found to be effective under anhydrous conditions because the alcohols were exchangeable with other organic solvents in solution and was more susceptible to lysozyme than to Chitinase, and showed and a poor susceptibility to chitosanase.
Abstract: Both the amount of water and the number of calcium ions are main factors affecting the dissolution of chitin in calcium chloride dihydrate-saturated methanol (calcium solvent). The higher degree of N-acetylation of the chitin was also indicated by its higher solubility in calcium solvent. The chitin hydrogel was prepared by adding a large excess of water to the chitin solution with vigorous stirring, followed by extensive dialysis against water or by filtration to remove the methanol and calcium ions. The water content of the chitin hydrogel was approximately 94–96% (w/v) and could be controlled by centrifugation. The chitin gel was also prepared by the addition of a large excess of alcohol, such as ethanol and iso-propanol, and these protocols were found to be effective under anhydrous conditions because the alcohols were exchangeable with other organic solvents in solution. The chitin hydrogel was more susceptible to lysozyme than to chitinase, and showed and a poor susceptibility to chitosanase. A α-chitin-type crystalline structure was regenerated from chitin sheets prepared from both α-chitin and β-chitin solutions in calcium solvent, but the β-chitin-type sheet was formed from the β-chitin hydrogel prepared by mechanical agitation in water. The α-chitin hydrogel solidified when thawed after freezing, but the β-chitin hydrogel prepared by mechanical agitation maintained its gel form even after prolonged freezing. Animal studies revealed a low toxicity for the chitin sheet and an acceleration of epidermal cell regeneration.
TL;DR: Chitin/SF blend nanofibrous matrix, particularly the one that contained 75% chitin and 25% SF, could be a potential candidate for tissue engineering scaffolds because it has both biomimetic three-dimensional structure and an excellent cell attachment and spreading for NHEK and NHEF.
TL;DR: This work shows that Knk is an apical GPI-linked protein that acts at the plasma membrane, and proposes a model in which Knk and the predicted chitin-binding protein Rtv form membrane complexes essential for epithelial tubulogenesis and cuticle formation through their specific role in directing chit in filament assembly.
Abstract: Precise epithelial tube diameters rely on coordinated cell shape changes and apical membrane enlargement during tube growth. Uniform tube expansion in the developing Drosophila trachea requires the assembly of a transient intraluminal chitin matrix, where chitin forms a broad cable that expands in accordance with lumen diameter growth. Like the chitinous procuticle, the tracheal luminal chitin cable displays a filamentous structure that presumably is important for matrix function. Here, we show that knickkopf (knk) and retroactive (rtv) are two new tube expansion mutants that fail to form filamentous chitin structures, both in the tracheal and cuticular chitin matrices. Mutations in knk and rtv are known to disrupt the embryonic cuticle, and our combined genetic analysis and chemical chitin inhibition experiments support the argument that Knk and Rtv specifically assist in chitin function. We show that Knk is an apical GPI-linked protein that acts at the plasma membrane. Subcellular mislocalization of Knk in previously identified tube expansion mutants that disrupt septate junction (SJ) proteins, further suggest that SJs promote chitinous matrix organization and uniform tube expansion by supporting polarized epithelial protein localization. We propose a model in which Knk and the predicted chitin-binding protein Rtv form membrane complexes essential for epithelial tubulogenesis and cuticle formation through their specific role in directing chitin filament assembly.
TL;DR: Bacillus subtilis W-118, a strain that produces antifungal materials, excreted a chitinase when cultured in a medium containing shrimp- and crab-shell powder as the major carbon source, which had the unique characteristics of a low molecular mass and acidic pI.
TL;DR: The available data for Serratia chitinases show that the chitinolytic machinery of this bacterium consists of two processive exo-enzymes that degrade the chITin chains in opposite directions (ChiA and ChiB) and a non-processive endo-enzyme, ChiC.
Abstract: Serratia marcescens produces three chitinases, ChiA, ChiB and ChiC which together enable the bacterium to efficiently degrade the insoluble chitin polymer We present an overview of the structural properties of these enzymes, as well as an analysis of their activities towards artificial chromogenic chito-oligosaccharide-based substrates, chito-oligosaccharides, chitin and chitosan We also present comparative inhibition data for the pseudotrisaccharide allosamidin (an analogue of the reaction intermediate) and the cyclic pentapeptide argadin The results show that the enzymes differ in terms of their subsite architecture and their efficiency towards chitinous substrates The idea that the three chitinases play different roles during chitin degradation was confirmed by the synergistic effects that were observed for certain combinations of the enzymes Studies of the degradation of the soluble heteropolymer chitosan provided insight into processivity Taken together, the available data for Serratia chitinas
TL;DR: The results indicate that the PGA/chitin blend nanofibrous matrix, particularly the one that contained 25% PGA and 75% chitin with bovine serum albumin coating, could be a good candidate for tissue engineering scaffolds, because it has an excellent cell attachment and spreading for normal human fibroblasts.
TL;DR: In this laboratory-scale study, chitin and chitosan flakes obtained from shrimp wastes as carrier material for a hydrocarbon-degrading bacterial strain showed the best performance during storage and seawater bioremediation.
TL;DR: It is shown that a homologous gene for chitin synthase is expressed in the shell forming tissue of larval Mytilus galloprovincialis even in early embryonic stages, the first clear‐cut indication for a functional role of cytoskeletal forces in the precisely controlled mineral deposition process of mollusk shell biogenesis.
TL;DR: This procedure is well suited to screening a large collection of yeast mutants or to evaluating effects of environmental conditions on cell wall polysaccharide content, and is also applicable to other fungal species, including Schizosaccharomyces pombe, Candida albicans and Aspergillus fumigatus.
Abstract: A simple and reliable method for quantitative determination of cell wall polymers in fungal cell with an s.e.m. of 5% is described. This protocol is based on the hydrolysis by sulfuric acid of beta-glucan, mannan, galactomannan and chitin present at different levels in the wall of yeasts and filamentous fungi into their corresponding monomers glucose, mannose, galactose and glucosamine. The released monosaccharides are subsequently separated and quantified by high-performance ionic chromatography coupled to pulse amperometry detection, with a detection limit of 1.0 mug ml(-1). This procedure is well suited to screening a large collection of yeast mutants or to evaluating effects of environmental conditions on cell wall polysaccharide content. This procedure is also applicable to other fungal species, including Schizosaccharomyces pombe, Candida albicans and Aspergillus fumigatus. Results can be obtained in 3 d.
TL;DR: The chitinase was characterized, and antifungal activity was observed against phytopathogens and the first 15 N-terminal amino-acid residues of the chit inase were determined.
Abstract: Streptomyces venezuelae P10 could produce extracellular chitinase in a medium containing 0.6% colloidal chitin that was fermented for 96 hours at 30°C. The enzyme was purified to apparent homogeneity with 80% saturation of ammonium sulfate as shown by chitin affinity chromatography and DEAE-cellulose anion-exchange chromatography. Sodium dodecyl sulfate–polyacrylamide gel electrophoresis (SDS-PAGE) of the enzyme showed a molecular weight of 66 kDa. The chitinase was characterized, and antifungal activity was observed against phytopathogens. Also, the first 15 N-terminal amino-acid residues of the chitinase were determined. The chitin hydrolysed products were N-acetylglucosamine and N, N’-diacetylchitobiose.
TL;DR: Experimental evidence is provided that lipids incorporated from the cuticle were the source of aliphatic polymer, which appears to account for most of the fossil record of terrestrial arthropod as well as marine arthropods that lacked a biomineralized exoskeleton.
Abstract: Modern arthropod cuticles consist of chitin fibres in a protein matrix, but those of fossil arthropods with an organic exoskeleton, particularly older than Tertiary, contain a dominant aliphatic component. This apparent contradiction was examined by subjecting modern cockroach, scorpion and shrimp cuticle to artificial maturation (350 °C/700 bars/24 h) following various chemical treatments, and analysing the products with pyrolysis–gas chromatography/mass spectrometry (Py–GC/MS). Analysis of artificially matured untreated cuticle yielded moieties related to phenols and alkylated substituents, pyridines, pyrroles and possibly indenes (derived from chitin). n -Alkyl amides, C16 and C18 fatty acids and alkane/alk-1-ene homologues ranging from C9 to C19 were also generated, the last indicating the presence of an n -alkyl component, similar in composition to that encountered in fossil arthropods. Similar pyrolysates were obtained from matured pure C16 and C18 fatty acids. Py–GC/MS of cuticles matured after lipid extraction and hydrolysis did not yield any aliphatic polymer. This provides direct experimental evidence that lipids incorporated from the cuticle were the source of aliphatic polymer. This process of in situ polymerization appears to account for most of the fossil record of terrestrial arthropods as well as marine arthropods that lacked a biomineralized exoskeleton.
TL;DR: Further studies are necessary to elucidate the relationship between reduced chitin synthesis and increased expression of AqCHS1 in order to shed new light on trafficking and regulation of chit in biosynthesis in the mosquito affected by diflubenzuron.
TL;DR: A frameshift mutation in chiA, encoding a chitinase, was validated for both strains, providing an explanation for the ascertained absence of chitinolytic activities and the concomitant possibility of producing highly viscous Chitin in a fermentational deproteinization process.
Abstract: Proteolytic but chitinase-deficient microbial cultures were isolated from shrimp shell waste and characterized. The most efficient isolate was found to be a mixed culture consisting of two Bacillus licheniformis strains, which were first determined microscopically and physiologically. Molecular characterization was carried out by sequencing the 16S rRNA gene of both strains. According to the residual protein and ash content, the chitin obtained by fermentation of such a mixed culture was found to be comparable to a commercially available, chemically processed product. However, the strikingly high viscosity (80 versus 10 mPa of the commercially available sample) indicates its superior quality. The two strains differed in colony morphology and in their secretion capabilities for degradative extracellular enzymes. Sequencing of the loci encoding amylase, cellulase, chitinases, and proteases, as well as the degS/degU operon, which is instrumental in the regulation of degradative enzymes, and the pga operon, which is responsible for polyglutamic acid production, revealed no differences. However, a frameshift mutation in chiA, encoding a chitinase, was validated for both strains, providing an explanation for the ascertained absence of chitinolytic activities and the concomitant possibility of producing highly viscous chitin in a fermentational deproteinization process.
TL;DR: Results show for the first time the involvement of genes encoding chitinases in prey-trap interaction and their differential expression and activity during prey trapping.
Abstract: The genus Nepenthes represents carnivorous plants with pitcher traps capable of efficient prey capture and digestion. The possible involvement of plant chitinases in this process was studied in Nepenthes khasiana. Two different types of endochitinases were identified in the liquid of closed traps exhibiting substrate specificity for either long chitin polymers or N-acetylglucosamine (GlcNAc) oligomers. Injection of chitin into such closed sterile pitchers induced the appearance of additional endochitinase isoenzymes, with substrate specificity only for long chitin polymers. No significant exochitinase (N-acetyl-b-glucosaminidase) or chitobiosidase activity could be detected in the non-induced or induced trap liquid. Four genes representing two subgroups of basic chitinases, denoted as Nkchit1b and Nkchit2b, were isolated from the secretory region of N. khasiana pitchers. The main differences between the two subgroups are the presence of a proline-rich hinge region only in NkCHIT1b and a C-terminal putative vacuole targeting extension only in NkCHIT2b, indicating different compartmentalization of the two enzymes. Reverse transcription–polymerase chain reaction (RT– PCR) evaluation of mRNA levels showed that the Nkchit2b genes are constitutively expressed in the secretory cells while transcription of Nkchit1b genes is induced by chitin injection. These results show for the first time the involvement of genes encoding chitinases in prey–trap interaction and their differential expression and activity during prey trapping.
TL;DR: Comparison of the extraction results of NAS with that from freshwater prawns indicated that more impurities were left in NAS chitin, suggesting that composition and structural arrangement of ch itin in shells influence the efficiency of ultrasound-assisted extraction.
Abstract: The influence of sonication during extraction of chitin from North Atlantic shrimp (NAS) shells (Pandalus borealis) on chitin yield, purity, and crystallinity was investigated. Shells were peeled, washed, lyophilized, ground, and suspended for 4 h in 0.25 M HCl (1: 40) at 40 degrees C followed by ultrasonication at 41 W/cm(2) for 0, 1, and 4 h, respectively. Demineralized shells were lyophilized, resuspended in 0.25 M NaOH (1: 40), and ultrasonicated at 41 W/cm(2) for 0, 1, and 4 h to remove proteins. The yield and mineral and protein contents were determined after each processing step. The purity of extracted chitin was determined from the total amount of glucosamine. The crystallinity index and size of crystals were calculated from wide-angle X-ray scattering measurements. Scanning electron microscope images were recorded to evaluate morphological changes in samples. The yield of chitin from NAS decreased from 16.5 to 11.4% for 0 and 1 h sonicated samples, respectively, which was attributed to increased concentrations of depolymerized materials in the wash water. Sonication did not enhance the removal of minerals. The application of ultrasound enhanced the removal of proteins from 39.8 to 10.6, 8.3, and 7.3% after 0, 1, and 4 h of sonication treatments. The crystallinity index of chitin decreased from 87.6 to 79.1 and 78.5% after 1 and 4 h of sonication, yielding chitosans with crystallinity indices of 76.7, 79.5, and 74.8% after deacetylation, respectively. Fourier transform infrared spectroscopy scans indicated that the degree of acetylation of chitins was unaffected by sonication. Comparison of the extraction results of NAS with that from freshwater prawns indicated that more impurities were left in NAS chitin, suggesting that composition and structural arrangement of chitin in shells influence the efficiency of ultrasound-assisted extraction.
TL;DR: Soil application of biocontrol agents in combination with chitin induced a significant increase in the activities of peroxidase (PO), polyphenol oxidase (PPO), phenylalanine ammonia-lyase (PAL), chit inase and b-1,3-glucanase in coconut palms infected with Ganoderma lucidum.
Abstract: The effect of soil application of biocontrol agents (Pseudomonas fluorescens, Trichoderma viride and T. harzianum) in combination with chitin on induction of phenolics and defense enzymes in coconut roots infected with Ganoderma lucidum, the causal agent of Ganoderma disease, was investigated. Soil application of these biocontrol formulations in combination with chitin induced a significant increase in the activities of peroxidase (PO), polyphenol oxidase (PPO), phenylalanine ammonia-lyase (PAL), chitinase and b-1,3-glucanase in the G. lucidum infected palms. Activities of both PAL and PO reached maximum levels within 3 d while the activity of PPO reached the maximum level 6 d after application of a mixture of P. fluorescens, T. viride and chitin. Isozyme analysis revealed that unique PO3 and PPO2 isozymes were induced in coconut palms treated with P. fluorescens + T. viride + chitin. Accumulation of phenolics was recorded 3 d after treatment and reached maximum levels 9 d after treatment application. Activity of chitinase was significantly increased from the third day after treatment imposition and continued to increase up to 9 to 12 d in all treatments. Chitinase isozyme analysis revealed that a unique Chit3 isoform was induced in coconut roots treated with P. fluorescens + T. viride + chitin. The b-1,3-glucanase activity was maximum 9 d after treatment application. The mechanisms by which P. fluorescens + T. viride + chitin reduced the incidence of Ganoderma disease in coconut may be related to its ability to induce defense mechanisms in coconut palms.
TL;DR: Although chitinase G was not capable of releasing 4‐methylumbelliferyl from artificial chitooligosaccharide substrates, it was capable of degrading longer chitoolsigOSaccharides at rates similar to those observed for other chit inases.
Abstract: We describe the cloning, overexpression, purification, characterization and crystal structure of chitinase G, a single-domain family 19 chitinase from the Gram-positive bacterium Streptomyces coelicolor A3(2). Although chitinase G was not capable of releasing 4-methylumbelliferyl from artificial chitooligosaccharide substrates, it was capable of degrading longer chitooligosaccharides at rates similar to those observed for other chitinases. The enzyme was also capable of degrading a colored colloidal chitin substrate (carboxymethyl-chitin-remazol-brilliant violet) and a small, presumably amorphous, subfraction of alpha-chitin and beta-chitin, but was not capable of degrading crystalline chitin completely. The crystal structures of chitinase G and a related Streptomyces chitinase, chitinase C [Kezuka Y, Ohishi M, Itoh Y, Watanabe J, Mitsutomi M, Watanabe T & Nonaka T (2006) J Mol Biol358, 472-484], showed that these bacterial family 19 chitinases lack several loops that extend the substrate-binding grooves in family 19 chitinases from plants. In accordance with these structural features, detailed analysis of the degradation of chitooligosaccharides by chitinase G showed that the enzyme has only four subsites (- 2 to + 2), as opposed to six (- 3 to + 3) for plant enzymes. The most prominent structural difference leading to reduced size of the substrate-binding groove is the deletion of a 13-residue loop between the two putatively catalytic glutamates. The importance of these two residues for catalysis was confirmed by a site-directed mutagenesis study.
TL;DR: This review emphasizes pharmaceutical applications of chitosan and its derivatives, and presents recent advances, and some therapeutical applications of these polymers are also discussed.
Abstract: Chitin and chitosan are copolymers build from N-acetyl-D-glucosamine and D-glucosamine. The former is widely found in nature and yields the latter on deacetylation. The copolymers are being used for several purposes. Since 1977, when the First International Conference on Chitin and Chitosan was held in Boston, USA, the interest on chitin and chitosan has remarkably increased. This review emphasizes pharmaceutical applications of chitosan and its derivatives, and presents recent advances. Some therapeutical applications of these polymers are also discussed.