In silico exploration of nitrogenase in Ensifer fredii unraveling structural homology functional and phylogenetic relationships and molecular dynamics

The nitrogenase Fe protein, central to nitrogen fixation, has been extensively studied, However, the X-ray structure of the Nase-Fe protein from Ensifer fredii is essential for understanding its function and stability, yet this structure is absent from the database. This study aimed to analyze the structural features of the NifH protein (EC 1.18.6.1, nitrogenase component II)) from Ensifer fredii using an in silico approach. Utilizing diverse biocomputational tools, we examined the primary, secondary, and tertiary structures of 22 NifH proteins, focusing on QSZ40499. Results indicate that QSZ40499 is a highly stable, hydrophilic, cytoplasmic protein with a molecular weight of 25.34 kDa. Its functional motif corresponds to the Fe4 family domain, and its secondary structure comprises 34.89% random coils and 34.47% alpha helices. 3D modeling via AlphaFold was validated using structural quality assessment servers and refined using the 3DRefiner server, supported by Z-scores and ERRAT Factor metrics. Molecular dynamics simulations revealed that QSZ40499 maintains structural stability and functional flexibility, essential for dynamic interactions during nitrogen fixation. This work provides valuable insights into the structural attributes of nitrogenase enzymes, advancing bioinformatics research and understanding of nitrogen fixation mechanisms.