Nitrogen (N) deposition influences microbial decomposition and nutrient transformation rates by affecting the activities of microbial properties, such as soil enzymes, microbial biomass and structure of the microbial community. In desert ecosystems, where water and N are two major limiting factors for biological activity, minimal nutrient dynamics are linked to comparatively large changes. In this paper, an in situ experiment was conducted in Gurbantunggut desert to explore the effects of N deposition on soil enzyme activities and microbial biomass N. Sixty 8 m × 8 m plots were established in the inter-dune area, with each plot having similar vegetation and a biological soil crust cover. Six N fertilizer treatments (N0, N0.5, N1, N3, N6 and N24 denoting 0, 0.5, 1.0, 3.0, 6.0 and 24.0 g N m^-2 a^-1, respectively) were conducted, with ten replicates for each treatment. Nitrogen was applied at the ratio of 2 ∶ 1 for NH⁺₄-N ∶ NO^-₃-N (informed in NH₄NO₃ and NH₄Cl), consisting with ratio of the main N components in deposition in Urumqi. Soil samples were collected in May 2009 from 24 of the 60 plots with 4 replicates for each treatment (6 N treatments × 4 replicates). Soil samples were taken from two layers (0-5 cm and 5-10 cm) and transported to laboratory quickly. The results showed that original vertical distribution patterns of nutrient, soil enzyme and microbial biomass were not changed by N addition. Soil organic C, total N, total P, available P and available N pools were greater in the 0-5 cm soil layer than in the 5-10 cm layer, regardless of N addition rates. The polyphenol oxidase and peroxidase activities in 0-5 cm soil layer were 14.3% - 27.3% and 1.4% - 14.2% lower than those in 5-10 cm soil layer, respectively. However, the activities of invertase, urease, alkaline phosphatase (AlP) and microbial biomass N in the 0-5 cm soil layer were 18.0% - 98.1%, 45.3% - 119.0%, 76.1% - 138.1% and 77.5% - 162.4% higher compared with the 5-10 cm layer, respectively. The soil enzyme activities and microbial biomass N were more sensitive to N addition in the 0-5 cm layer than in the 5-10 cm layer. Although low and moderate N levels (N0.5, N1 and N3) have marginal effects on oxidative enzyme activities in the 0-5 cm layer, high N levels (N6 and N24) reduced the activities of oxidative enzyme. With respect to control (N0), the activities of polyphenol oxidase and peroxidase were 22.4% and 12.1% lower under N24. In contrast, the oxidative enzyme activities in the 5-10 cm layer were not sensitive to N addition. With the increase in N addition, both the invertase and AlP activities in the two layers increased at low N addition levels, then they decreased at high N levels. Compared with control, N1 and N3 increased invertase activities in the 0-5 cm layer by 49.6% and 72.3%, respectively; and by 8.6% and 11.4% in the 5-10 cm layer, respectively. AlP activities in the 0-5 cm layer tended to increase at the low N levels (i.e., N0.5, N1 and N3). Highest increasing rate of 48.6% was found in N3 treatment in comparison to control. Nitrogen additions resulted in increased microbial biomass N, but decreased urease activities. With the changes in soil enzyme activities, cycling of C, N and P altered accordingly. The varying soil organic C under different N treatments may be partially due to the change of oxidative activities. The addition of N lead to the increase of microbial biomass N and available N. Available P increased significantly at low N addition rates while it decreased at high N addition rates in both the soil layers. Our results suggest that N addition or deposition can affect the nutrient flow by changing the soil enzyme activities and microbial biomass in Gurbantunggut desert ecosystem.