8peb

OXA-48_Q5. Epistasis Arises from Shifting the Rate-Limiting Step during Enzyme Evolution

spinqualitylabelsall models 8peb, resolution 1.17Å Show:Asymmetric Unit Biological Assembly Drag the structure with the mouse to rotate   Export Animated Image   Print3D Structural highlights 8peb is a 1 chain structure with sequence from Klebsiella pneumoniae. Full crystallographic information is available from OCA. For a guided tour on the structure components use FirstGlance. Method: X-ray diffraction, Resolution 1.17Å Ligands: CL, MLI, PEG, SO4 Resources: FirstGlance, OCA, PDBe, RCSB, PDBsum, ProSAT Publication Abstract from PubMed Epistasis, the non-additive effect of mutations, can provide combinatorial improvements to enzyme activity that substantially exceed the gains from individual mutations. Yet the molecular mechanisms of epistasis remain elusive, undermining our ability to predict pathogen evolution and engineer biocatalysts. Here we reveal how directed evolution of a beta-lactamase yielded highly epistatic activity enhancements. Evolution selected four mutations that increase antibiotic resistance 40-fold, despite their marginal individual effects (</=2-fold). Synergistic improvements coincided with the introduction of super-stochiometric burst kinetics, indicating that epistasis is rooted in the enzyme's conformational dynamics. Our analysis reveals that epistasis stemmed from distinct effects of each mutation on the catalytic cycle. The initial mutation increased protein flexibility and accelerated substrate binding, which is rate-limiting in the wild-type enzyme. Subsequent mutations predominantly boosted the chemical steps by fine-tuning substrate interactions. Our work identifies an overlooked cause for epistasis: changing the rate-limiting step can result in substantial synergy that boosts enzyme activity. Epistasis arises from shifting the rate-limiting step during enzyme evolution of a beta-lactamase.,Frohlich C, Bunzel HA, Buda K, Mulholland AJ, van der Kamp MW, Johnsen PJ, Leiros HS, Tokuriki N Nat Catal. 2024;7(5):499-509. doi: 10.1038/s41929-024-01117-4. Epub 2024 Feb 23. PMID:38828429 From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine.

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