9ezd

BsmI (Bottom Nicking mutant) crystallized with Mg2+ and cognate dsDNA (Post-reactive complex)

spinqualitylabelsall models 9ezd, resolution 2.63Å Show:Asymmetric Unit Biological Assembly Drag the structure with the mouse to rotate   Export Animated Image   Print3D Structural highlights 9ezd is a 4 chain structure with sequence from Geobacillus stearothermophilus. Full crystallographic information is available from OCA. For a guided tour on the structure components use FirstGlance. Method: X-ray diffraction, Resolution 2.635Å Ligands: CL, MES, MG Resources: FirstGlance, OCA, PDBe, RCSB, PDBsum, ProSAT Function Q8RLN4_GEOSE Publication Abstract from PubMed BsmI, a thermophilic Type IIS restriction endonuclease from Bacillus stearothermophilus, presents a unique structural composition, housing two distinct active sites within a single monomer. Recognition of the non-symmetrical 5'-GAATGC-3' sequence enables precise cleavage of the top and bottom DNA strands. Synthetic biology interventions have led to the transformation of BsmI into Nb.BsmI, a nicking endonuclease. Here we introduce Nt*.BsmI, tailored for top-strand cleavage, which is inactive on standard double-stranded DNA, but active on bottom-strand nicked DNA, suggesting a sequential cleavage mechanism. Crystallographic structures of pre- and post-reactive complexes with cognate DNA show one major conformational change, a retractable loop possibly governing sequential active site accessibility. The x-ray structures reveal the position of the divalent metal ions in the active sites and the DNA:protein interactions, while the models predicted by Alphafold3 are incorrect. This comprehensive structural and functional study lays a foundation for rational enzyme redesign and potential applications in biotechnology. Crystal structures of monomeric BsmI restriction endonuclease reveal coordinated sequential cleavage of two DNA strands.,Sieskind R, Missoury S, Madru C, Commenge I, Niogret G, Hollenstein M, Rondelez Y, Sauguet L, Haouz A, Legrand P, Delarue M Commun Biol. 2025 Mar 7;8(1):387. doi: 10.1038/s42003-025-07612-z. PMID:40055548 From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine.

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