Abstract

Recently, a crystal structure of V-nitrogenase was presented, showing that one of the µ₂ sulphide ions in the active site (S2B) is replaced by a lighter atom, suggested to be NH or NH₂, i.e. representing a reaction intermediate. Moreover, a sulphur atom is found 7 Å from the S2B site, suggested to represent a storage site for this ion when it is displaced. We have re-evaluated this structure with quantum refinement, i.e. standard crystallographic refinement in which the empirical restraints (employed to ensure that the final structure makes chemical sense) are replaced by more accurate quantum–mechanical calculations. This allows us to test various interpretations of the structure, employing quantum–mechanical calculations to predict the ideal structure and to use crystallographic measures like the real-space Z-score and electron-density difference maps to decide which structure fits the crystallographic raw data best. We show that the structure contains an OH⁻-bound state, rather than an N₂-derived reaction intermediate. Moreover, the structure shows dual conformations in the active site with ~ 14% undissociated S2B ligand, but the storage site seems to be fully occupied, weakening the suggestion that it represents a storage site for the dissociated ligand.

Graphic abstract

中文翻译：

---

钒固氮酶的晶体结构是否包含反应中间体？来自量子细化的证据。

摘要

最近，提出了V-氮酶的晶体结构，表明活性位点（S2B）中的µ ₂硫化物离子之一被较轻的原子取代，建议是NH或NH ₂，即代表反应中间体。此外，在S2B位点发现7Å的硫原子，表示该离子被置换时代表该离子的储存位点。我们用量子细化重新评估了这种结构，即标准的晶体学细化，其中经验约束（用于确保最终结构具有化学意义）被更精确的量子力学计算所代替。这使我们能够测试结构的各种解释，采用量子力学计算来预测理想结构，并使用诸如实空间Z分数和电子密度差图之类的晶体学方法来确定哪种结构最适合晶体学原始数据。我们表明，该结构包含OH ^-结合态，而不是N ₂衍生的反应中间体。此外，该结构在活性位点显示约14％未解离的S2B配体的双重构象，但该存储位点似乎被完全占据，从而削弱了它代表解离的配体的存储位点的暗示。

图形摘要