Metagenomics and bio-engineering of chitin and chitosan modifying enzymes for biotechnological applications

Chitin ist nach Cellulose das am häufigsten in der Natur vorkommende Biopolymer. Chitosan ist die vollständige oder teilweise deacetylierte Version des Chitins. Chitin und Chitosan modifizierende Enzyme (CCME) finden als Werkzeuge zur Erzeugung spezieller Chitosane für die Biotechnologie und Biomedizin Anwendung. In dieser Arbeit wurde sowohl mithilfe einer metagenomischen Genbank als auch mittels mikrobieller Direktisolaten nach neuen CCMEs gesucht. Als Ausgangsmaterial für diese beiden Ansätze dienten Bodenproben, die über mehr als zehn Jahre in Kontakt mit Chitin und Chitosan waren. CCME kodierende Gene aus Bacillus spp. wurden in E. coli-Stämmen heterolog exprimiert und die entsprechenden Enzyme aufgereinigt. Eine Inkubation von Chitosanpolymeren verschiedener Acetylierungsgrade (50%, 35% und 10%) mit den aufgereinigten Enzymen generierte Mischungen aus Chitin- und Chitosan-Oligomeren, die vielversprechende Elicitor- und Priming-Effekte auf pflanzliche Zellkulturen hatten

Chitosan encapsulation modulates the effect of capsaicin on the tight junctions of MDCK cells

Capsaicin has known pharmacological effects including the ability to reversibly open cellular tight junctions, among others. The aim of this study was to develop a strategy to enhance the paracellular transport of a substance with low permeability (FITC-dextran) across an epithelial cell monolayer via reversible opening of cellular tight junctions using a nanosystem comprised by capsaicin and of chitosan. We compared the biophysical properties of free capsaicin and capsaicin-loaded chitosan nanocapsules, including their cytotoxicity towards epithelial MDCK-C7 cells and their effect on the integrity of tight junctions, membrane permeability and cellular uptake. The cytotoxic response of MDCK-C7 cells to capsaicin at a concentration of 500 μM, which was evident for the free compound, is not observable following its encapsulation. The interaction between nanocapsules and the tight junctions of MDCK-C7 cells was investigated by impedance spectroscopy, digital holographic microscopy and structured illumination fluorescence microscopy. The nanocapsules modulated the interaction between capsaicin and tight junctions as shown by the different time profile of trans-epithelial electrical resistance and the enhanced permeability of monolayers incubated with FITC-dextran. Structured illumination fluorescence microscopy showed that the nanocapsules were internalized by MDCK-C7 cells. The capsaicin-loaded nanocapsules could be further developed as drug nanocarriers with enhanced epithelial permeability

Ergatis: a web interface and scalable software system for bioinformatics workflows

Motivation: The growth of sequence data has been accompanied by an increasing need to analyze data on distributed computer clusters. The use of these systems for routine analysis requires scalable and robust software for data management of large datasets. Software is also needed to simplify data management and make large-scale bioinformatics analysis accessible and reproducible to a wide class of target users

A High Diversity in Chitinolytic and Chitosanolytic Species and Enzymes and Their Oligomeric Products Exist in Soil with a History of Chitin and Chitosan Exposure

Chitin is one of the most abundant biomolecules on earth, and its partially de-N-acetylated counterpart, chitosan, is one of the most promising biotechnological resources due to its diversity in structure and function. Recently, chitin and chitosan modifying enzymes (CCMEs) have gained increasing interest as tools to engineer chitosans with specific functions and reliable performance in biotechnological and biomedical applications. In a search for novel CCME, we isolated chitinolytic and chitosanolytic microorganisms from soils with more than ten-years history of chitin and chitosan exposure and screened them for chitinase and chitosanase isoenzymes as well as for their patterns of oligomeric products by incubating their secretomes with chitosan polymers. Of the 60 bacterial strains isolated, only eight were chitinolytic and/or chitosanolytic, while 20 out of 25 fungal isolates were chitinolytic and/or chitosanolytic. The bacterial isolates produced rather similar patterns of chitinolytic and chitosanolytic enzymes, while the fungal isolates produced a much broader range of different isoenzymes. Furthermore, diverse mixtures of oligosaccharides were formed when chitosan polymers were incubated with the secretomes of select fungal species. Our study indicates that soils with a history of chitin and chitosan exposure are a good source of novel CCME for chitosan bioengineering