X‐Ray Crystallographic Analysis of NifB with a Full Complement of Clusters: Structural Insights into the Radical SAM‐Dependent Carbide Insertion During Nitrogenase Cofactor Assembly

• Published in: Angewandte Chemie International Edition 2021-02, Vol.60 (5), p.2364-2370
• Main Authors: Kang, Wonchull, Rettberg, Lee A., Stiebritz, Martin T., Jasniewski, Andrew J., Tanifuji, Kazuki, Lee, Chi Chung, Ribbe, Markus W., Hu, Yilin
• Format: Article
• Language: English
• Subjects: Assembly; Biosynthesis; carbide insertion; Carbides; Clusters; cofactors; Crystal structure; Crystallography; Crystallography, X-Ray - methods; Insertion; Models, Molecular; Molecular Structure; Nitrogenase; Nitrogenase - chemistry; nitrogenases; radical SAM enzyme; S-Adenosylmethionine - chemistry; structural biology; Substrates
• ISSN: 1433-7851; 1521-3773
• DOI: 10.1002/anie.202011367

NifB is an essential radical SAM enzyme required for the assembly of an 8Fe core of the nitrogenase cofactor. Herein, we report the X‐ray crystal structures of Methanobacterium thermoautotrophicum NifB without (apo MtNifB) and with (holo MtNifB) a full complement of three [Fe4S4] clusters. Both apo and holo MtNifB contain a partial TIM barrel core, but unlike apo MtNifB, holo MtNifB is fully assembled and competent in cofactor biosynthesis. The radical SAM (RS)‐cluster is coordinated by three Cys, and the adjacent K1‐ and K2‐clusters, representing the precursor to an 8Fe cofactor core, are each coordinated by one His and two Cys. Prediction of substrate channels, combined with in silico docking of SAM in holo MtNifB, suggests the binding of SAM between the RS‐ and K2‐clusters and putative paths for entry of SAM and exit of products of SAM cleavage, thereby providing important mechanistic insights into the radical SAM‐dependent carbide insertion concomitant with cofactor core formation. NifB is an essential radical SAM enzyme required for the assembly of an 8Fe core of the nitrogenase cofactor. Here we report the crystal structures of Methanobacterium thermoautotrophicum NifB without and with a full complement of three [Fe4S4] clusters, which provide important mechanistic insights into the radical SAM‐dependent carbide insertion concomitant with the formation of a cofactor core.