The continuously increasing concentration of atmospheric CO₂ has considerably altered ecosystem functioning. However, few studies have examined the long-term (i.e. over a decade) effect of elevated CO₂ on soil microbial communities. Using 16S rRNA gene amplicons and a GeoChip microarray, we investigated soil microbial communities from a Californian annual grassland after 14 years of experimentally elevated CO₂ (275 ppm higher than ambient). Both taxonomic and functional gene compositions of the soil microbial community were modified by elevated CO₂. There was decrease in relative abundance for taxa with higher ribosomal RNA operon (rrn) copy number under elevated CO₂, which is a functional trait that responds positively to resource availability in culture. In contrast, taxa with lower rrn copy number were increased by elevated CO₂. As a consequence, the abundance-weighted average rrn copy number of significantly changed OTUs declined from 2.27 at ambient CO₂ to 2.01 at elevated CO₂. The nitrogen (N) fixation gene nifH and the ammonium-oxidizing gene amoA significantly decreased under elevated CO₂ by 12.6% and 6.1%, respectively. Concomitantly, nitrifying enzyme activity decreased by 48.3% under elevated CO₂, albeit this change was not significant. There was also a substantial but insignificant decrease in available soil N, with both nitrate (NO₃⁻) (−27.4%) and ammonium (NH₄⁺) (−15.4%) declining. Further, a large number of microbial genes related to carbon (C) degradation were also affected by elevated CO₂, whereas those related to C fixation remained largely unchanged. The overall changes in microbial communities and soil N pools induced by long-term elevated CO₂ suggest constrained microbial N decomposition, thereby slowing the potential maximum growth rate of the microbial community.