Biochemical characterization of recombinant L-threonine 3-dehydrogenase from Akkermansia muciniphila

L-threonine is an essential amino acid required for mucin synthesis and intestinal barrier maintenance. The gut commensal Akkermansia muciniphila, which resides in the mucus layer, encodes a putative L-threonine 3-dehydrogenase (Am_TDH) whose biochemical properties remain unknown. Here, we report the heterologous expression, purification, and biochemical characterization of this enzyme. The enzyme was characterized as a member of the zinc-containing medium-chain dehydrogenase/reductase (MDR) superfamily and shown to form a homotetramer with a molecular mass of 154 kDa. Am_TDH was strictly NAD⁺-dependent, exhibiting maximal specific activity toward L-threonine (30.3 ± 0.4 U·mg^-1). Optimal catalytic activity was observed at pH 8.5 and 37 °C. The K_m values for L-threonine and NAD⁺ at 37 °C were 3.56 ± 0.48 mM and 0.65 ± 0.19 mM, respectively. These findings reveal a physiologically adapted, stereospecific TDH and provide molecular insights into amino acid metabolism by a gut symbiont.