l-Arabinitol 4-dehydrogenase (LAD) from Hypocrea jecorina (HjLAD) was cloned and overexpressed in Escherichia coli BL21 (DE3). The kinetics of l-arabinitol oxidation by NAD(+), catalyzed by HjLAD, was studied within the pH range of 7.0-9.5 at 25°C. The turnover number (kcat) and the catalytic efficiency (kcat/Km) were 4200min(-1) and 290mM(-1)min(-1), respectively. HjLAD showed the highest turnover number and catalytic efficiency among all previously characterized LADs. In further application of HjLAD, rare l-sugar l-xylulose was produced by the enzymatic oxidation of arabinitol to give a yield of approximately 86%.