Looking at Nitrogenase: Insights from Modern Structural Approaches

Abstract

Nitrogenase, the primary biological source of fixed nitrogen, has been studied by various biochemical and biophysical methods to determine the mechanism of nitrogen reduction to ammonia. Previously, structural studies have contributed to determining the arrangement and identity of the unique metallocofactors of the as-isolated nitrogenase enzyme. Due to the multi-protein, dynamic nature of catalysis in nitrogenase, structurally capturing intermediates is not trivial. Recently, we have developed methods for preparing crystallographic samples of nitrogenase from active assay mixtures. The "out-of-assay" approach has yielded structures of small molecules bound to the active site cofactor, revealing an unexpected rearrangement of the belt sulfur atoms. The activity-based methods provide a framework for accessing non-resting states of the cofactor and introduce new questions surrounding the controlled binding and release of substrates. In the following, we discuss recent structural advances in the field and the novel directions for future activity-based research.

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