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| Thimoteo,S.S.; Glogauer,A.; Faoro,H.; de Souza,E.M.; Huergo,L.F.; Moerschbacher,B.M.; Pedrosa,F.O.. |
| Chitinases are hydrolases that degrade chitin, a polymer of N-acetylglucosamine linked β(1-4) present in the exoskeleton of crustaceans, insects, nematodes and fungal cell walls. A metagenome fosmid library from a wastewater-contaminated soil was functionally screened for chitinase activity leading to the isolation and identification of a chitinase gene named metachi18A. The metachi18A gene was subcloned and overexpressed in Escherichia coli BL21 and the MetaChi18A chitinase was purified by affinity chromatography as a 6xHis-tagged fusion protein. The MetaChi18A enzyme is a 92-kDa protein with a conserved active site domain of glycosyl hydrolases family 18. It hydrolyses colloidal chitin with an optimum pH of 5 and temperature of 50°C. Moreover, the enzyme... |
| Tipo: Info:eu-repo/semantics/article |
Palavras-chave: Chitinase; Metagenomic; Aeromonas; Kinetics. |
| Ano: 2017 |
URL: http://www.scielo.br/scielo.php?script=sci_arttext&pid=S0100-879X2017000100602 |
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| Alves,L.P.S.; Almeida,A.T.; Cruz,L.M.; Pedrosa,F.O.; de Souza,E.M.; Chubatsu,L.S.; Müller-Santos,M.; Valdameri,G.. |
| The conventional method for quantification of polyhydroxyalkanoates based on whole-cell methanolysis and gas chromatography (GC) is laborious and time-consuming. In this work, a method based on flow cytometry of Nile red stained bacterial cells was established to quantify poly-3-hydroxybutyrate (PHB) production by the diazotrophic and plant-associated bacteria, Herbaspirillum seropedicae and Azospirillum brasilense. The method consists of three steps: i) cell permeabilization, ii) Nile red staining, and iii) analysis by flow cytometry. The method was optimized step-by-step and can be carried out in less than 5 min. The final results indicated a high correlation coefficient (R2=0.99) compared to a standard method based on methanolysis and GC. This method... |
| Tipo: Info:eu-repo/semantics/article |
Palavras-chave: Flow cytometry; Nile red; Poly-3-hydroxybutyrate; Herbaspirillum seropedicae; Azospirillum brasilense. |
| Ano: 2017 |
URL: http://www.scielo.br/scielo.php?script=sci_arttext&pid=S0100-879X2017000100606 |
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