A bacterial tungsten-containing aldehyde oxidoreductase forms an enzymatic decorated protein nanowire

Aldehyde oxidoreductases (AORs) are tungsten enzymes catalyzing the oxidation of many different aldehydes to the corresponding carboxylic acids. In contrast to other known AORs, the enzyme from the denitrifying betaproteobacterium Aromatoleum aromaticum (AORAa) consists of three different subunits (AorABC) and uses nicotinamide adenine dinucleotide (NAD) as an electron acceptor. Here, we reveal that the enzyme forms filaments of repeating AorAB protomers that are capped by a single NAD-binding AorC subunit, based on solving its structure via cryo-electron microscopy. The polyferredoxin-like subunit AorA oligomerizes to an electronconducting nanowire that is decorated with enzymatically active and W-cofactor (W-co) containing AorB subunits. Our structure further reveals the binding mode of the native substrate benzoate in the AorB active site. This, together with quantum mechanics:molecular mechanics (QM:MM)-based modeling for the coordination of the W-co, enables formulation of a hypothetical catalytic mechanism that paves the way to further engineering for applications in synthetic biology and biotechnology.

Automated summary

In this study, the cryo-electron microscopy technique was used to reveal that the aldehyde oxidoreductases (AORs) from Aromatoleum aromaticum form filaments consisting of repeating AorAB protomers capped by a single NAD-binding AorC subunit. The structure also reveals the binding mode of the native substrate benzoate and the electronconducting nanowire formed by the polyferredoxin-like subunit AorA. A hypothetical catalytic mechanism was proposed based on quantum mechanics:molecular mechanics (QM:MM) modeling, which provides insights for further engineering in synthetic biology and biotechnology applications.