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Dynamic coordination of two-metal-ions orchestrates λ-exonuclease catalysis
Metal ions at the active site of an enzyme act as cofactors, and their dynamic fluctuations can potentially influence enzyme activity. Here, we use λ-exonuclease as a model enzyme with two Mg(2+) binding sites and probe activity at various concentrations of magnesium by single-molecule-FRET. We find...
Autores principales: | , , , , , , , , , , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
Nature Publishing Group UK
2018
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6199318/ https://www.ncbi.nlm.nih.gov/pubmed/30353000 http://dx.doi.org/10.1038/s41467-018-06750-9 |
Sumario: | Metal ions at the active site of an enzyme act as cofactors, and their dynamic fluctuations can potentially influence enzyme activity. Here, we use λ-exonuclease as a model enzyme with two Mg(2+) binding sites and probe activity at various concentrations of magnesium by single-molecule-FRET. We find that while Mg(A)(2+) and Mg(B)(2+) have similar binding constants, the dissociation rate of Mg(A)(2+) is two order of magnitude lower than that of Mg(B)(2+) due to a kinetic-barrier-difference. At physiological Mg(2+) concentration, the Mg(B)(2+) ion near the 5’-terminal side of the scissile phosphate dissociates each-round of degradation, facilitating a series of DNA cleavages via fast product-release concomitant with enzyme-translocation. At a low magnesium concentration, occasional dissociation and slow re-coordination of Mg(A)(2+) result in pauses during processive degradation. Our study highlights the importance of metal-ion-coordination dynamics in correlation with the enzymatic reaction-steps, and offers insights into the origin of dynamic heterogeneity in enzymatic catalysis. |
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