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Upon infection by pathogenic bacteria, production of reactive oxygen species (ROS) is part of the host organism's first line of defence. ROS damage a number of macromolecules, and in order to withstand such a harsh environment, the bacteria need to have well‐functioning ROS scavenging and repair systems. Herein, MutT is an important nucleotide‐pool sanitization enzyme, which degrades 8‐oxo‐dGTP and thus prevents it from being incorporated into DNA. In this context, we have performed a comparative biochemical and structural analysis of MutT from the fish pathogenAliivibrio salmonicida (AsMutT) and the human pathogenVibrio cholerae (VcMutT), in order to analyse their function as nucleotide sanitization enzymes and also determine possible cold‐adapted properties ofAsMutT. The biochemical characterisation revealed that both enzymes possess activity towards the 8‐oxo‐dGTP substrate, and thatAsMutT has a higher catalytic efficiency thanVcMutT at all temperatures studied. Calculations based on the biochemical data also revealed a lower activation energy (Ea) forAsMutT compared toVcMutT, and differential scanning calorimetry experiments showed thatAsMutT displayed an unexpected higher melting temperature (Tm) value thanVcMutT. A comparative analysis of the crystal structure ofVcMutT, determined to 2.42 Å resolution, and homology models ofAsMutT indicate that three unique Gly residues in loops ofVcMutT, and additional long range ion‐pairs inAsMutT could explain the difference in temperature stability of the two enzymes. We conclude thatAsMutT is a stable, cold‐active enzyme with high catalytic efficiency and reducedEa, compared to the mesophilicVcMutT.