Abstract
Peptidoglycan (PG) is a highly cross-linked, protective mesh-like sacculus that surrounds the bacterial cytoplasmic membrane. Expansion of PG is tightly coupled to growth of a bacterial cell and requires hydrolases to cleave the cross-links for insertion of nascent PG material. In Escherichia coli, a proteolytic system comprising the periplasmic PDZ-protease Prc and the lipoprotein adaptor NlpI contributes to PG enlargement by regulating cellular levels of MepS, a cross-link-specific hydrolase. Here, we demonstrate how NlpI binds Prc to facilitate the degradation of its substrate MepS by structural and mutational analyses. An NlpI homodimer binds two molecules of Prc and forms three-sided MepS-docking cradles using its tetratricopeptide repeats. Prc forms a monomeric bowl-shaped structure with a lid-like PDZ domain connected by a substrate-sensing hinge that recognizes the bound C terminus of the substrate. In summary, our study reveals mechanistic details of protein degradation by the PDZ-protease Prc bound to its cognate adaptor protein.
Highlights
Peptidoglycan (PG) is a highly cross-linked, protective mesh-like sacculus that surrounds the bacterial cytoplasmic membrane
The levels of MepS are regulated via proteolysis by a newly identified protease complex consisting of Prc, a soluble periplasmic PDZ-protease, and NlpI, an outer membrane (OM) lipoprotein with tetratricopeptide repeats (TPRs)[3, 4]
The purified sNlpI-Prc complex predominantly formed a 2:2 tetramer (Fig. 1a, b), with a determined Mw of 239.8 ± 24.6 kDa based on Analytical ultracentrifugation (AUC) and 230.2 ± 8.7 kDa based on size exclusion chromatography with multi-angle static light scattering (SEC-MALS)
Summary
Peptidoglycan (PG) is a highly cross-linked, protective mesh-like sacculus that surrounds the bacterial cytoplasmic membrane. In Escherichia coli, a proteolytic system comprising the periplasmic PDZ-protease Prc and the lipoprotein adaptor NlpI contributes to PG enlargement by regulating cellular levels of MepS, a cross-link-specific hydrolase. The NlpI-Prc system completely degrades MepS into small fragments[3], which is unlike the other wellknown C-terminal processing PDZ proteases that cleave the C termini of specific precursor protein substrates[5, 6]. Prc belongs to the large family of C-terminal processing proteases that recognize the C terminus of protein substrates[7]. Many of these PDZ-containing proteases, such as CtpB and D1P, are mostly involved in cleaving the C termini of specific protein substrates[5, 6].
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