The cobalamin processing enzyme of Trichoplax adhaerens.

Abstract

Cobalamin (Cbl) is an essential cofactor for methionine synthase (MS) and methylmalonyl-CoA mutase (MUT), but it must first undergo chemical processing for utilization in animals. In humans, this processing comprises β-axial ligand cleavage and Cbl reduction and is performed by the enzyme MMACHC (HsCblC). Although the functionality of CblC is well-understood in higher order organisms, little is known about the evolutionary origin of these enzymes and the reactivity of CblCs in lower-order organisms with unique environmental and cellular conditions. Therefore, we investigated the CblC of Trichoplax adhaerens (TaCblC), a marine organism considered to be one of the earliest evolutionarily diverging and simplest living animals. The TaCblC sequence contained conserved residues important for Cbl processing in higher-order organisms. The predicted structure of TaCblC closely resembled known CblC structures and had features consistent with Cbl and co-substrate binding capabilities. Recombinantly expressed TaCblC could bind and process several Cbl analogues using glutathione or NADH as co-substrates, similarly to previously characterized CblCs, but with variable rates and dependencies on the presence of oxygen. Notably, TaCblC dealkylates methylcobalamin at a rate ca. 2-times higher than HsCblC, although this comes with a lower ratio of product to glutathione oxidation, suggesting higher unproductive electron transfer in the TaCblC system. This reflects differences in cellular conditions of the more ancient homologue, which lives in low oxygen levels and an environment of low Cbl biovailability (∼2 pM in sea water).