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Dissecting the link between the enzymatic activity and the SaPI inducing capacity of the phage 80α dUTPase

DOI: 10.1038/s41598-017-11234-9 DOI Help

Authors: Christian Alite (Instituto de Biomedicina de Valencia (IBV-CSIC); CIBER de Enfermedades Raras (CIBERER)) , Suzanne Humphrey (University of Glasgow) , Jordi Donderis (Instituto de Biomedicina de Valencia (IBV-CSIC); CIBER de Enfermedades Raras (CIBERER)) , Elisa Maiques (Instituto de Biomedicina de Valencia (IBV-CSIC); CIBER de Enfermedades Raras (CIBERER)) , J. Rafael Ciges-tomas (Instituto de Biomedicina de Valencia (IBV-CSIC); CIBER de Enfermedades Raras (CIBERER)) , José R. Penadés (University of Glasgow) , Alberto Marina (Instituto de Biomedicina de Valencia (IBV-CSIC); CIBER de Enfermedades Raras (CIBERER))
Co-authored by industrial partner: No

Type: Journal Paper
Journal: Scientific Reports , VOL 7

State: Published (Approved)
Published: September 2017
Diamond Proposal Number(s): 14739

Open Access Open Access

Abstract: The trimeric staphylococcal phage-encoded dUTPases (Duts) are signalling molecules that induce the cycle of some Staphylococcal pathogenicity islands (SaPIs) by binding to the SaPI-encoded Stl repressor. To perform this regulatory role, these Duts require an extra motif VI, as well as the Dut conserved motifs IV and V. While the apo form of Dut is required for the interaction with the Stl repressor, usually only those Duts with normal enzymatic activity can induce the SaPI cycle. To understand the link between the enzymatic activities and inducing capacities of the Dut protein, we analysed the structural, biochemical and physiological characteristics of the Dut80α D95E mutant, which loses the SaPI cycle induction capacity despite retaining enzymatic activity. Asp95 is located at the threefold central channel of the trimeric Dut where it chelates a divalent ion. Here, using state-of-the-art techniques, we demonstrate that D95E mutation has an epistatic effect on the motifs involved in Stl binding. Thus, ion binding in the central channel correlates with the capacity of motif V to twist and order in the SaPI-inducing disposition, while the tip of motif VI is disturbed. These alterations in turn reduce the affinity for the Stl repressor and the capacity to induce the SaPI cycle.

Journal Keywords: Bacteriophages; Enzyme mechanisms; Phage biology; X-ray crystallography

Subject Areas: Biology and Bio-materials, Medicine


Instruments: I04-Macromolecular Crystallography

Other Facilities: ALBA

Documents:
s41598-017-11234-9.pdf