Centre d'Étude et de Valorisation de la Diversité Microbienne, Département de Biologie, Faculté des Sciences, Université de Sherbrooke, 2500 boulevard de l'Université, Sherbrooke, J1K 2R1, (Québec) Canada.
Microb Cell Fact. 2011 Feb 10;10:7. doi: 10.1186/1475-2859-10-7.
Chitosanases 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.
Two 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.
This 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.
壳聚糖酶是一种水解壳聚糖的酶，壳聚糖是一种β-1,4 连接的 D-葡萄糖胺生物聚合物。壳寡糖具有许多新兴的应用，壳聚糖酶可用于壳聚糖的工业酶解。这些细胞外酶由许多生物体产生，包括真菌和细菌，在生化和酶学水平上得到了很好的研究，但很少有工作致力于它们的基因表达调控。这是第一个关于链霉菌中产异源壳聚糖酶基因（csnN106）的遗传调控的研究。
两个链霉菌菌株用于诱导实验：野生型菌株及其突变体（ΔcsnR），其 csnR 基因发生了框内缺失，编码一个负转录调节因子。在各种培养基中比较壳聚糖酶水平表明，CsnR 负调控异源壳聚糖酶基因 csnN106 的表达。使用 ΔcsnR 宿主和一个经过修饰的转录操作子的突变 csnN106 基因，可以在没有壳聚糖的情况下，使用廉价的培养基成分大量生产壳聚糖酶。此外，由于观察到较低水平的细胞外蛋白酶和蛋白质污染物，壳聚糖酶的产量质量更高。
这种新的壳聚糖酶生产系统具有生物技术意义，因为仅使用普通的培养基成分，并直接从培养上清液中获得高纯度的酶。
