Climate change and soil acidification are critical factors affecting crop production and soil quality. This study comprehensively analyzed the impact of fertilization practices, including conventional (CA), sustainable (SA), and unfertilized (BK), on soil properties, enzyme activities, and gene diversity in paddy fields across seasonal changes. Soil pH was significantly influenced by fertilization, with higher pH in BK and a decrease in pH with increased fertilization. Soil enzyme activities and Biolog EcoPlate™ analysis revealed the lowest activities in September, with the highest in December under different practices. Metagenomic analysis showed the highest genetic richness in CA soil, with seasonal variations influencing genetic diversity. From the perspective of genes in species taxonomy, Sorangium cellulosum and Anaeromyxobacter sp. were the most abundant taxa. Soil genes annotated by CAZy, COG, and GO databases revealed highly similar gene structures among different practices. Moreover, the genetic origins of soil enzymes were linked to specific bacterial contributors. While not all gene’s diversity and abundance were associated with soil enzyme activity, arylsulfatase showed an obvious correlation. Enzyme activities proved more sensitive indicators of microbial activity than gene abundance. This study emphasizes the need for rational fertilization strategies to maintain soil enzyme activities, considering agricultural practices and seasonal variations.