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Vegetation and soil microorganisms are important components of terrestrial ecosystems and play a crucial role in ecosystem functioning. However, little is known about the synergistic changes in soil microbial community with aboveground plants in grassland degradation and the role of the microbial community in the process of vegetation restoration succession. In this study, we investigated the characteristics of soil microbial communities and diversities in the different levels of grassland degradation using Illumina MiSeq high-throughput sequencing. The dominant bacteria phyla were: Actinobacteriota, 31.61–48.90%; Acidobacteriota, 7.19–21.73%; Chloroflexi, 9.08–19.09%; and Proteobacteria, 11.14–18.03%. While the dominant fungi phyla were: Ascomycota, 46.36–81.58%; Basidiomycota, 5.63–33.18%; and Mortierellomycota, 1.52–37.69%. Through RDA/CCA, the effects of environmental factors on the differences in the soil microbial community between different sites were interpreted. Results showed that the pH was the most critical factor affecting soil microbial communities in seriously degraded grassland; nevertheless, soil microbial communities in non-degraded grassland and less degraded grasslands were mainly affected by the soil moisture content and soil enzyme activities (sucrase activity, alkaline phosphatase activity and catalase activity). We systematically demonstrated the soil microbial communities of different grassland degradation gradients in Mongolia, which provided valuable information for grassland degradation reduction and vegetation restoration succession.