Abstract: In this study, the β-xylosidase gene xlyY411, belonging to glycoside hydrolase family 39 (GH39), was mined from the metagenome of a hot spring in Yuanjiang County, Yunnan Province, China. The gene was heterologously expressed in Escherichia coli BL21 and purer recombinant enzyme XlyY411 was obtained by affinity chromatography. The optimum temperature of XlyY411 was 85 ℃, and the t_1/2 at 90 ℃ was 6.2 h. The optimum pH was 5.0, and there was a good pH tolerance between pH 5−9 at 4 ℃. XlyY411 displayed a good tolerance to surfactants, while its activity of was above 75% with the addition of 10% β-mercaptoethanol or tween; XlyY411 displayed a good tolerance to organic solvents, while its activity of was above 50% with the addition of 20% methanol, ethanol or isopropanol; There was 51.6% enzyme activity at a xylose concentration of 2 mmol/L, showing its good product tolerance. Enzymatic kinetic studies showed that XlyY411 had a K_M of (7.19 ± 0.89) mmol, a v_max of 4753.68 mmol/(mg·min), a k_cat of 223.91 s, and a k_cat/K_M of 31.14 s⁻¹·mmol⁻¹. The molecular docking analysis indicate that XlyY411 had similar catalytic mechanism to the β-xylosidases reported in literature.