Regulation of a Phage Endolysin by Disulfide Caging

In contrast to canonical phage endolysins, which require holin-mediated disruption of the membrane to gain access to attack the cell wall, signal anchor release (SAR) endolysins are secreted by the hostsec system, where they accumulate in an inactive form tethered to the membrane by their N-terminal SAR domains. SAR endolysins become activated by various mechanisms upon release from the membrane. In its inactive form, the prototype SAR endolysin, LyzP1 , of coliphage P1, has an active-site Cys covalently blocked by a disulfide bond; activation involves a disulfide bond isomerization driven by a thiol in the newly released SAR domain, unblocking the active-site Cys. Here, we report that Lyz103 , the endolysin ofErwinia phage ERA103, is also a SAR endolysin. Although Lyz103 does not have a catalytic Cys, genetic evidence suggests that it also is activated by a thiol-disulfide isomerization triggered by a thiol in the SAR domain. In this case, the inhibitory disulfide in nascent Lyz103 is formed between cysteine residues flanking a catalytic glutamate, caging the active site. Thus, LyzP1 and Lyz103 define subclasses of SAR endolysins that differ in the nature of their inhibitory disulfide, and Lyz103 is the first enzyme found to be regulated by disulfide bond caging of its active site.