The crystal structure of the major pneumococcal autolysin LytA in complex with a large peptidoglycan fragment reveals the pivotal role of glycans for lytic activity

Abstract

The pneumococcal autolysin LytA is a key virulence factor involved in several important functions including DNA competence, immune evasion and biofilm formation. Here, we present the 1.05 Å crystal structure of the catalytic domain of LytA in complex with a synthetic cell-wall-based peptidoglycan (PG) ligand that occupies the entire Y-shaped substrate-binding crevice. As many as twenty-one amino-acid residues are engaged in ligand interactions with a majority of these interactions directed towards the glycan strand. All saccharides are intimately bound through hydrogen bond, van der Waals and CH-π interactions. Importantly, the structure of LytA is not altered upon ligand binding, whereas the bound ligand assumes a different conformation compared to the unbound NMR-based solution structure of the same PG-fragment. Mutational study reveals that several non-catalytic glycan-interacting residues, structurally conserved in other amidases from Gram-positive Firmicutes, are pivotal for enzymatic activity. The three-dimensional structure of the LytA/PG complex provides a novel structural basis for ligand restriction by the pneumococcal autolysin, revealing for the first time an importance of the multivalent binding to PG saccharides.

Bacterial size matters: Multiple mechanisms controlling septum cleavage and diplococcus formation are critical for the virulence of the opportunistic pathogen Enterococcus faecalis

“…A substrate-assisted catalytic mechanism involving anchimeric assistance by the C2-acetamido group of the GlcNAc moiety is likely to underpin this requirement [ 25 ]. Another example is the pneumococcal autolysin LytA, in which several amino acids in the vicinity of the catalytic residues contribute to positioning of the substrate in the catalytic cleft so that the scissile bond is at an optimal distance from the catalytic residue [ 26 ]. Our work revealed that the N -acetylglucosaminidase domain of AtlA is not essential for septum cleavage.…”

ClyJ Is a Novel Pneumococcal Chimeric Lysin with a Cysteine- and Histidine-Dependent Amidohydrolase/Peptidase Catalytic Domain

“…LytA is the major autolysin of S. pneumoniae essential for bacterial survival and cell wall synthesis (23). Recent structural elucidation of the full-length LytA and its complex with large peptidoglycan fragments reveals new insights into its function and lysis mechanism (24,25), providing new clues for developing chimeric lysins against S. pneumoniae. PlyC, a multimeric streptococcal lysin composed of one PlyCA subunit and eight subunits of PlyCB, is specific for groups A, C, and E streptococci (26)(27)(28).…”

Computationally Aided Discovery of LysEFm5 Variants with Improved Catalytic Activity and Stability

“…The catalytic domain of the major pneumococcal autolysin LytA, initially evaluated due to the availability of the solved crystal structure of the domain bound to a synthetic peptidoglycan ligand (37), was identified as a homolog of the catalytic domain of LysEFm5 via sequence alignment (with a sequence similarity of 0.23). SWISS-Model provided a QMEAN score of Ϫ7.58, sequence identity of 8.33, and coverage of 1.00 when the catalytic domain of LytA (PDB code 5CTV) was used as a template to model the first 180 amino acids in the catalytic domain of LysEFm5.…”

“…N83 in LysEFm5, the only putative primary residue considered in this analysis, is structurally analogous to N79 in LytA. N79 is a ligand-binding residue that is highly conserved across multiple prokaryote-and eukaryote-derived peptidoglycan recognition proteins (PGRPs), both with and without amidase activity (37). In AmiE (the amidase domain of the major autolysin of Staphylococcus epidermidis) and in human PGRP-I␣, this conserved asparagine residue was shown to hydrogen bond with the carbonyl groups in the second and third amino acids in the peptide stem of N-acetylmuramic acid (MurNAc)-L-Ala-D-isoGln-L-Lys, a peptidoglycan analog (23,46).…”