Abstract
BackgroundIn the previous study, the cellulolytic Escherichia coli ZH-4 isolated from bovine rumen was found to show extracellular cellulase activity and could degrade cellulose in the culture. The goal of this work was to identify and characterize the secreted cellulase of E. coli ZH-4. It will be helpful to re-understand E. coli and extend its application in industry.ResultsA secreted cellulase was confirmed to be endo-glucanase BcsZ which was encoded by bcsZ gene and located in the cellulose synthase operon bcsABZC in cellulolytic E. coli ZH-4 by western blotting. Characterization of BcsZ indicated that a broad range of pH and temperature tolerance with optima at pH 6.0 and 50 °C, respectively. The apparent Michaelis–Menten constant (Km) and maximal reaction rate (Vmax) for BcsZ were 8.86 mg/mL and 0.3 μM/min·mg, respectively. Enzyme activity of BcsZ was enhanced by Mg2+ and inhibited by Zn2+, Cu2+ and Fe3+. BcsZ could hydrolyze carboxymethylcellulose (CMC) to produce cello-oligosaccharides, cellotriose, cellobiose and glucose.ConclusionsIt is confirmed that extracellular cellulolytic capability of E. coli ZH-4 was attributed to BcsZ, which explained why E. coli ZH-4 can grow on cellulose. The endo-glucanase BcsZ from E. coli-ZH4 has some new characteristics which will extend the understanding of endo-glucanase. Analysis of the secretion characteristics of BcsZ provided a great reference for applying E. coli in multiple industrial fields.
Highlights
In the previous study, the cellulolytic Escherichia coli ZH-4 isolated from bovine rumen was found to show extracellular cellulase activity and could degrade cellulose in the culture
BcsZ was detected in culture medium (Fig. 3a, line 1) with the recombinant cell by Sodium dodecyl sulfate polyacrylamide gel Identification and verification of secreted cellulase in cellulolytic E. coli ZH-4 The mature protein in the culture medium was analyzed through western blotting to identify the extracellular endoglucanase in ZH-4
Sequence analysis showed that the promoter, Fnr and FNR-binding site of bcsZ in E. coli ZH-4 is consistent with MG1655 and W3110
Summary
The cellulolytic Escherichia coli ZH-4 isolated from bovine rumen was found to show extracellular cellulase activity and could degrade cellulose in the culture. The goal of this work was to identify and characterize the secreted cellulase of E. coli ZH-4. Cellulose biomass is the most abundant carbohydrate on the earth. Cellulose as the main component of plant cell wall consists of linear long chains of β-1, 4 glucose units. Hydrolyzing cellulose by cellulase is ideal and promising for its utilization in environmentally friendly and high efficiency manner [2]. The cooperative action of three kinds of cellulolytic enzymes (endo-glucanase, exo-glucanase, and β-glucosidase) is essential in hydrolysis of cellulose to glucose [3]. Among three kinds of cellulolytic enzymes, endoglucanases plays an important role in
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