Extended Data Fig. 6: Mhp from diverse staphylococcal phages are sensed by Sau–Thoeris.
From: Bacterial TIR-based immune systems sense phage capsids to initiate defense
(a) Growth of S. aureus RN4220 harboring plasmids carrying either an incomplete (thsB1/B2) or full (thsA/B1/B2) Thoeris operon in the presence of a second plasmid expressing Mhp from different staphylococcal phages, determined as the OD600 of the cultures after addition of IPTG. The values at the end of the curves, at 16 hours, were used to make the bar graphs shown in Fig. 6a. Mean of +/- S.D. of three biological replicates is reported. (b) Coomassie Blue-stained SDS-PAGE of Mhp proteins, purified from staphylococci harboring pMhp plasmids using PEG-enrichment. Protein molecular weight (kDa) markers are shown. (c) HPLC analysis of the products resulting from the incubation of purified His6-ThsA with the products of the reactions shown in Fig. 6c, obtained after mixing ThsB2-His6, ThsB2F6A-His6, NAD+ and purified Mhp from different staphylococcal phages. Retention times (RT) of reactants and products are marked by dotted lines. (d) Phylogenetic tree of the staphylococcal Mhps used in this study, generated using EMBL-EBI Muscle multiple sequence alignment tool. The branch numbers indicate genetic distance from ΦNM1 Mhp calculated by the software. (e) Alphafold3 prediction of the monomeric form of each Mhp encoded by the different phages. Structural distance is shown as the RMSD of the different Mhps compared to ΦNM1 Mhp (pTMs: Φ12γ3 pTM = 0.76, ΦNM4 pTM = 0.80, Φ80α pTM = 0.79, ΦJ1 pTM = 0.78, ΦNM1 pTM = .75, ΦJ4 pTM = 0.81).
