Lei_2021_Microb.Cell.Fact_20_197

BACKGROUND: Herbaspirillum camelliae is a gram-negative endophyte isolated from the tea plant. Both strains WT00C and WT00F were found to hydrolyze epigallocatechin-3-gallate (EGCG) and epicatechin-3-gallate (ECG) to release gallic acid (GA) and display tannase activity. However, no tannase gene was annotated in the genome of H. camelliae WT00C. RESULTS: The 39 kDa protein, annotated as the prolyl oligopeptidase in the NCBI database, was finally identified as a novel tannase. Its gene was cloned, and the enzyme was expressed in E. coli and purified to homogeneity. Moreover, enzymatic characterizations of this novel tannase named Tan(Hcw) were studied. Tan(Hcw) was a secretary enzyme with a Sec/SPI signal peptide of 48 amino acids at the N-terminus, and it catalyzed the degradation of tannin, methyl gallate (MG), epigallocatechin-3-gallate (EGCG) and epicatechin-3-gallate (ECG). The optimal temperature and pH of Tan(Hcw) activities were 30 degreesC, pH 6.0 for MG and 40 degreesC, pH 7.0 for both EGCG and ECG. Na(+), K(+) Mn(2+) and Triton-X100, Tween80 increased the enzyme activity of Tan(Hcw), whereas Zn(2+), Mg(2+), Hg(2+), EMSO, EDTA and beta-mercaptoethanol inhibited enzyme activity. K(m), k(cat) and k(cat) /K(m) of Tan(Hcw) were 0.30 mM, 37.84 s(-1), 130.67 mM(-1) s(-1) for EGCG, 0.33 mM, 34.59 s(-1), 105.01 mM(-1) s(-1) for ECG and 0.82 mM, 14.64 s(-1), 18.17 mM(-1) s(-1) for MG, respectively. CONCLUSION: A novel tannase Tan(Hcw) from H. camelliae has been identified and characterized. The biological properties of Tan(Hcw) suggest that it plays a crucial role in the specific colonization of H. camelliae in tea plants. Discovery of the tannase Tan(Hcw) in this study gives us a reasonable explanation for the host specificity of H. camelliae. In addition, studying the characteristics of this enzyme offers the possibility of further defining its potential in industrial application.