Heavy metals such as arsenic (As), lead (Pb), copper (Cu) and zinc (Zn) can be found in large concentrations in mine spills in Mexico. Interest in contamination by these heavy metals has increased recently as they can change the functioning of soil ecosystems qualitatively and quantitatively. They disturb the activities of soil fauna and contaminate drinking water in large parts of the world, which severely affects human health. Little, however, is known how heavy metals might affect the biological functioning of a soil. Soil was sampled from eight locations along a gradient of heavy-metal contamination with distance from a mine in San Luis Potosí (Mexico) active since about 1800 AD. Microbial biomass was determined with the original chloroform fumigation incubation (CFI) as well as extraction (CFE) techniques and the substrate induced respiration (SIR) technique while C and N mineralization were measured. Total concentrations of As in the top 0–10 cm soil layer ranged from 8 to 22992 mg kg–1, from 31 to 1845 mg kg–1 for Pb, from 27 to 1620 mg kg–1 for Cu and from 81 to 4218 mg kg–1 for Zn. There was a significant negative correlation (P < 0.0001) between microbial biomass, soil organic carbon, total N and C mineralization and the heavy metal content of the soil. The microbial biomass C to organic C ratio, which varied from 0.4 to 1.9%, specific respiratory activity (qCO2), and oxidation of NO2– were not affected by heavy metals. It was found that long-term contamination of soil with heavy metals had an adverse effect on the amount of soil microorganisms as evidenced by a marked decrease in microbial biomass C, but not some of their characteristics. According to principal components analysis (PCA), the correlation matrix showed three distinct factors explaining 71% of the variance. A first factor including heavy metals (As, Pb, Cu and Zn) with a negative loading and total N, organic C, soil microbial biomass with a positive loading characterized the soil organic matter and contamination status. Loam and sand combined for the second factor characterizing the textural classification while the third factor was loaded by CEC and clay content.