Breaking Benzene Aromaticity—Computational Insights into the Mechanism of the Tungsten-Containing Benzoyl-CoA Reductase

Abstract

Aromatic compounds are environmental pollutants with toxic and carcinogenic properties. Despite the stability of aromatic rings, bacteria are able to degrade the aromatic compounds into simple metabolites and use them as growth substrates under oxic or even under anoxic conditions. In anaerobic microorganisms, most monocyclic aromatic growth substrates are converted to the central intermediate benzoyl-coenzyme A, which is enzymatically reduced to cyclohexa-1,5-dienoyl-CoA. The strictly anaerobic bacterium Geobacter metallireducens uses the class II benzoyl-CoA reductase complex for this reaction. The catalytic BamB subunit of this complex harbors an active site tungsten-bis-pyranopterin cofactor with the metal being coordinated by five protein/cofactor-derived sulfur atoms and a sixth, so far unknown, ligand. Although BamB has been biochemically and structurally characterized, its mechanism still remains elusive. Here we use continuum electrostatic and QM/MM calculations to model benzoyl-CoA reduction by BamB. We aim to elucidate the identity of the sixth ligand of the active-site tungsten ion together with the interplay of the electron and proton transfer events during the aromatic ring reduction. On the basis of our calculations, we propose that benzoyl-CoA reduction is initiated by a hydrogen atom transfer from a W(IV) species with an aqua ligand, yielding W(V)-[OH] and a substrate radical intermediate. In the next step, a proton-assisted second electron transfer takes place with a conserved active-site histidine serving as the second proton donor. Interestingly, our calculations suggest that the electron for the second reduction step is taken from the pyranopterin cofactors rather than from the tungsten ion. The resulting cationic radical, which is distributed over both pyranopterins, is stabilized by conserved anionic amino acid residues. The stepwise mechanism of the reduction shows similarities to the Birch reduction known from organic chemistry. However, the strict coupling of protons and electrons allows the reaction to proceed under milder conditions.

Computational Understanding of the Selectivities in Metalloenzymes

“…The mechanism and selectivity of three different tungstoenzymes have been subjected to QM and QM/MM calculations, namely, acetylene hydratase (Liao et al, 2010; Liao and Himo, 2011), formaldehyde oxidoreductase (Liao et al, 2011b; Liao, 2013) and benzoyl CoA reductase (Culka et al, 2017; Qian and Liao, 2018).…”

Facile Benzene Reduction by a Ca2+/AlI Lewis Acid/Base Combination

“…Three possible routes for formation of (BDI)Ca + (C 6 H 6 )Al III -(BDI). [22] Pathway 2s tarts with dissociation of (BDI)Ca + ·(C 6 H 6 ) into the fragments (BDI)Ca + and C 6 H 6 .T his reaction is endergonic by + 18.1 kcal mol À1 ,avalue which is too high because the counteranion that stabilizes (BDI)Ca + better than (BDI)Ca + ·(C 6 H 6 )i sn eglected. The counteranion was neglectedb ut the full BDI ligand, CH[C(CH 3 )N-Dipp] 2 ,h as been included, except for the model system for route 1s hown at the top.…”

“…[21] Nature makes use of as imilar proton assisted reduction that breaks down the aromaticity of an arene by simultaneous proton and electron transfer from aW IV (H 2 O)/W V (OH) couple (Scheme 3). [22] Pathway 2s tarts with dissociation of (BDI)Ca + ·(C 6 H 6 ) into the fragments (BDI)Ca + and C 6 H 6 .T his reaction is endergonic by + 18.1 kcal mol À1 ,avalue which is too high because the counteranion that stabilizes (BDI)Ca + better than (BDI)Ca + ·(C 6 H 6 )i sn eglected. Thec alculated energies, however, nicely show that coordination of (BDI)Al I is much more exergonic (À38.3 kcal mol À1 )t han that of benzene (À18.1 kcal mol À1 ).…”

One-megadalton metalloenzyme complex in Geobacter metallireducens involved in benzene ring reduction beyond the biological redox window

“…reduction (20,21). However, the components and mechanisms of the underlying energetic coupling have remained hypothetical.…”