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Urbanization in the Arctic results in considerable and still poorly known environmental consequences. The effect of urbanization on soil microbiome—an ecosystem component highly sensi-tive to anthropogenic disturbance—remains overlooked for the Arctic region. The research compared chemical and microbial properties of the natural Podzol soils and urban soils of Murmansk—the largest Arctic city. Particular attention was given to the profile distribution, which is almost com-pletely ignored by most microbial studies. Soil microbiome was investigated by the quantitative indicators based on fluorescence microscopy (microbial biomass) and PCR real-time methods (amount of rRNA genes copies of archaea, bacteria, and fungi). The principal changes in urban soils’ properties compared to the natural references included a shift in pH and an increase in C and nutrients’ contents, especially remarkable for the subsoil. The numbers of rRNA genes copies of archaea, bacteria, and fungi in urban topsoils (106 –1010, 109 –1010, and 107 –109, respectively) were lower than in Podzol; however, the opposite pattern was shown for the subsoil. Similarly, the total microbial biomass in urban topsoils (0.55–0.75 mg g−1 ) was lower compared to the 1.02 mg g−1 in Podzols, while urban subsoil microbial biomass was 2–2.5 times higher than in the natural conditions. Both for urban and natural soils and throughout the profiles, fungi were dominated by mycelium forms; however, the ratios of mycelium–spores were lower, and the amount of thin mycelium was higher in urban soils than in natural Podzols. Urbanization in the Arctic altered soil morphological and chemical properties and created a new niche for microbial development in urban subsoils; its contribution to biodiversity and nutrient cycling promises to become increasingly important under projected climate change.