Deciphering how Cpl-7 cell wall-binding repeats recognize the bacterial peptidoglycan

Abstract

Endolysins, the cell wall lytic enzymes encoded by bacteriophages to release the phage progeny, are among the top alternatives to fight against multiresistant pathogenic bacteria; one of the current biggest challenges to global health. Their narrow range of susceptible bacteria relies, primarily, on targeting specific cell-wall receptors through specialized modules. The cell wall-binding domain of Cpl-7 endolysin, made of three CW_7 repeats, accounts for its extended-range of substrates. Using as model system the cell wall-binding domain of Cpl-7, here we describe the molecular basis for the bacterial cell wall recognition by the CW_7 motif, which is widely represented in sequences of cell wall hydrolases. We report the crystal and solution structure of the full-length domain, identify N-acetyl-D-glucosaminyl-(β1,4)-N-acetylmuramyl-L-alanyl-D-isoglutamine (GMDP) as the peptidoglycan (PG) target recognized by the CW_7 motifs, and characterize feasible GMDP-CW_7 contacts. Our data suggest that Cpl-7 cell wall-binding domain might simultaneously bind to three PG chains, and also highlight the potential use of CW_7-containing lysins as novel anti-infectives.

Structural and functional insights into a novel two-component endolysin encoded by a single gene in Enterococcus faecalis phage

“…Catalytic domains are responsible for cleavage of peptidoglycan bonds within the cellwall and can be classified into four groups: N-acetylglucosaminidases, N-acetylmuramoyl-Lalanine amidases, N-acetylmuramidases (lysozymes), and endopeptidases [18][19][20]. In contrast, CBDs display much greater variety and distinguish discrete epitopes present within the host cell-wall, typically carbohydrates or teichoic acids, thus giving rise to species-specific or strainspecific activity of a particular endolysin [21,22]. The modular structure of lysins makes it possible to design bioengineered endolysins that have desired properties, such as higher activity, or broader killing spectrum.…”

Catalytic diversity and cell wall binding repeats in the phage‐encoded endolysins

“…CBDs composed of a three‐helix bundle is present in both Gram‐positive and modular Gram‐negative phage endolysins (Table ). The Gram‐positive targeting example, Cpl‐7, uses three repeats of the three helix bundle CW_7 fold to bind to N ‐acetyl‐ d ‐glucosaminyl‐(β1,4)‐ N ‐acetylmuramyl‐ l ‐alanyl‐ d ‐isoglutamine in PG (Bustamante et al ., ) (Fig. C).…”

“…Computational docking combined with saturation‐transfer difference nuclear magnetic resonance spectroscopy suggest that N ‐acetyl‐ d ‐glucosaminyl‐(β1,4)‐ N ‐acetylmuramyl‐ l ‐alanyl‐ d ‐isoglutamine binds in a shallow groove located between the two last helices of the bundle and the first one at the bottom (Bustamante et al ., ). The interaction is mediated by residues that are conserved in the CW_7 family (Bustamante et al ., ), suggesting other CW_7 domains may bind similar target ligands. The S. agalactiae phage endolysin, λSA2 possesses two CW_7 repeats in an unusual central position between an endopeptidase (N‐terminal) and a glucosaminidase (C‐terminal) domain (Pritchard et al ., ).…”

Microarray Strategies for Exploring Bacterial Surface Glycans and Their Interactions With Glycan-Binding Proteins

“…An example is the study of the recognition of the bacterial peptidoglycan by the cell wall-binding domain of the endolysin Cpl-7, which is encoded by the pneumococcal Cp-7 bacteriophage (Bustamante et al, 2017). Cpl-7 is composed of a catalytic domain with muramidase activity and a cell wall-binding domain (C-Cpl-7) made up of three CW_7 repeats.…”