Allosteric regulation of DegS protease subunits though a shared energy landscape

Abstract

The PDZ domains of the trimeric DegS protease bind unassembled outer-membrane proteins (OMPs) that accumulate in the E. coli periplasm. This cooperative binding reaction triggers a proteolytic cascade that activates a transcriptional stress response. To dissect the mechanism of allosteric activation, we generated hybrid DegS trimers with different numbers of PDZ domains and/or protease-domain mutations. By studying the chemical reactivity and enzymatic properties of these hybrids, we show that all subunits experience a strongly coupled energetic landscape. For example, OMP-peptide binding to a single PDZ domain stimulates active-site chemical modification and proteolytic cleavage in the attached and neighboring protease domains. OMP-peptide binding relieves inhibitory PDZ interactions, whereas the interfaces between protease domains in the trimeric DegS core mediate positively cooperative activation driven both by substrate binding and inhibition relief.