Why some nitrogen-processing enzymes are more efficient than others

Nitrogen, a prevalent gas in Earth's atmosphere, remains inaccessible to most life forms. A few microbes, equipped with nitrogenases, can break nitrogen molecules apart and transform them into ammonia. These nitrogenases come in three varieties, distinguished by the metals they incorporate. Among these, nitrogenases laden with molybdenum demonstrate the highest efficiency.

Two recent studies from MIT shed light on the reasons behind this superior efficiency, which could aid in creating engineered or synthetic catalysts capable of converting nitrogen gas to ammonia. Molybdenum, although not directly binding to nitrogen, facilitates iron atoms in locking onto nitrogen more robustly. This initial binding is a crucial hurdle, but once broken, the subsequent steps become relatively straightforward.

Prior to the evolution of nitrogen-fixing abilities, around 3 billion years ago, even high-energy phenomena like lightning were insufficient to separate nitrogen atoms. The emergence of nitrogenase enzymes altered this situation, enabling cells to extract nitrogen from the air for biomass. Within the active site of nitrogenase lies a catalytic cofactor comprising iron, sulfur, carbon, and sometimes another metal.

Among nitrogenases, those with molybdenum in their cofactors exhibit superior efficiency, followed by those with vanadium, and the least efficient are those with only iron. The reason for this efficiency gap has puzzled scientists, particularly given that molybdenum allegedly does not bind directly to nitrogen gas. To unravel this mystery, MIT researchers developed simplified versions of iron-sulfur clusters found in naturally occurring cofactors.

These clusters, modifiable by incorporating different metal atoms, allowed them to study how varying metals impact cofactor properties. In the first paper, led by Wu and Ehweiner, the team substituted various metal atoms and measured the iron's ability to bind nitrogen. They discovered that only cofactors containing large metal atoms, such as molybdenum or tungsten, could strongly grasp N2.

Conversely, vanadium, chromium, or iron, being smaller, did not bind N2 and engaged in alternate reactions. This paper mirrors the biological observations that iron-sulfur clusters harboring molybdenum are better at binding dinitrogen than those with lighter metals. The second paper, spearheaded by Brown, delves into a potential explanation for this phenomenon.

Here, the researchers examined how cofactors with different metals interact with N-heterocyclic carbenes. These molecules behave akin to N2 in certain aspects and serve as a suitable model for this investigation. Like N2, they resist accepting electrons from another molecule, a critical step in breaking chemical bonds. The investigation revealed that molybdenum's presence within the cluster simplifies iron's electron donation to N-heterocyclic carbenes through a process called back-bonding.

This occurs because molybdenum, being a large atom, possesses extensive orbitals that can overlap with the orbitals of the nearby iron atom. This alteration in iron's electron density, facilitated by molybdenum's interaction, makes it easier for iron to transfer electrons to N2. Once N2 binds to an iron atom, the remaining reaction can transpire smoothly.

A proton can then enter from water or another source to create an N-H bond, which subsequently simplifies the subsequent breaking and binding of N-N bonds to protons, culminating in the formation of ammonia. These findings could guide researchers in engineering enzymes for organisms to generate their own NH3, potentially reducing or eliminating the need for artificial fertilizers.

Additionally, they could assist chemists in devising synthetic catalysts that produce ammonia industrially while consuming less energy than the Haber–Bosch process. The overarching implication of these findings is to provide insights into nature's approach to this vital and remarkable reaction, potentially translating into new processes.