Abstract
BackgroundChitosanases are enzymes hydrolysing chitosan, a β-1,4 linked D-glucosamine bio-polymer. Chitosan oligosaccharides have numerous emerging applications and chitosanases can be used for industrial enzymatic hydrolysis of chitosan. These extracellular enzymes, produced by many organisms including fungi and bacteria, are well studied at the biochemical and enzymatic level but very few works were dedicated to the regulation of their gene expression. This is the first study on the genetic regulation of a heterologous chitosanase gene (csnN106) in Streptomyces lividans.ResultsTwo S. lividans strains were used for induction experiments: the wild type strain and its mutant (ΔcsnR), harbouring an in-frame deletion of the csnR gene, encoding a negative transcriptional regulator. Comparison of chitosanase levels in various media indicated that CsnR regulates negatively the expression of the heterologous chitosanase gene csnN106. Using the ΔcsnR host and a mutated csnN106 gene with a modified transcription operator, substantial levels of chitosanase could be produced in the absence of chitosan, using inexpensive medium components. Furthermore, chitosanase production was of higher quality as lower levels of extracellular protease and protein contaminants were observed.ConclusionsThis new chitosanase production system is of interest for biotechnology as only common media components are used and enzyme of high degree of purity is obtained directly in the culture supernatant.
Highlights
Chitosanases are enzymes hydrolysing chitosan, a b-1,4 linked D-glucosamine bio-polymer
While these enzymes are endo-hydrolases, their mechanism could potentially be transformed into exo-type by protein engineering as shown for the GH46 chitosanase from Bacillus circulans MH-K1 [25]
Similar sequences are present upstream from the coding sequences of chitosanase genes found in other genomes of actinomycetes, displaying a clear consensus (Figure 2B)
Summary
Chitosanases are enzymes hydrolysing chitosan, a b-1,4 linked D-glucosamine bio-polymer. Chitosan oligosaccharides have numerous emerging applications and chitosanases can be used for industrial enzymatic hydrolysis of chitosan These extracellular enzymes, produced by many organisms including fungi and bacteria, are well studied at the biochemical and enzymatic level but very few works were dedicated to the regulation of their gene expression. This is the first study on the genetic regulation of a heterologous chitosanase gene (csnN106) in Streptomyces lividans. Chitosan is produced at industrial scale by alkaline deacetylation of chitin, originating mainly from crustacean shells [4] This polysaccharide, almost unique among natural polymers for its amino groups that remain positively charged in mild acidic solutions, is the subject of numerous works oriented towards its. Most studies were limited to the follow up of chitosanase production in various culture media [30,31]
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