FIG 2.

phiKT gp28 is necessary and sufficient for OM disruption during phage lysis. (A) E. coli 4s cultures (squares) were monitored for lysis after infection at t = 0 min with phiKT at an MOI of ∼5 in the presence (open circles) or absence (filled circles) of 10 mM Mg²⁺ supplementation. (Inset) A single cell imaged at the time of lysis, with frames spanning approximately 1 s total (5 μm scale bar). (B) E. coli MG1655 cells carrying plasmids encoding full or partial phage lysis cassettes were induced by the addition of 1 mM IPTG at t = 0 min. Each construct carries the plasmid pQ, with lambda Q under lacPO control, and various derivatives of pRE, where lysis genes from phiKT or lambda are under the control of the lambda pR’ (Q-dependent) promoter. pRE inserts: none, squares (“vector”); phiKT 27-28-29, filled circles (“28⁺”); phiKT 27-Δ28-29, open circles with dotted line (“Δ28”); 27-28_E17X-29, open circles with dashed line (“28_am”); lambda S105-R-Rz-Rz1, filled diamonds (“Rz⁺Rz1⁺”); lambda S105-R-Rz_am-Rz1_am, open diamonds (“Rz_amRz1_am”). A single cell imaged at t = 60 min is shown for vector, Rz_amRz1_am, and Δ28 with a 5 μm scale bar. (C) Cells carrying the spanin-defective prophage λ900 Rz_amRz1_am, pQ, and various pRE constructs were induced by thermal induction and the addition of 1 mM IPTG at t = 0 min. The pRE plasmid constructs are labeled identically as in panel B. (D) Identical to panel C, including thermal induction at t = 0 min except the prophage is λhy21(SRRz_amRz1_am)²¹ and encodes the phage 21 pinholin and SAR endolysin. The “28⁺” curves were induced for gp28 expression at t = 0 or 15 min, as indicated. All lysis curves were performed at least three times with similar results. A representative graph is shown.