Multiligand Specificity of Pathogen-associated Molecular Pattern-binding Site in Peptidoglycan Recognition Protein

Abstract

The peptidoglycan recognition protein PGRP-S is an innate immunity molecule that specifically interacts with microbial peptidoglycans and other pathogen-associated molecular patterns. We report here two structures of the unique tetrameric camel PGRP-S (CPGRP-S) complexed with (i) muramyl dipeptide (MDP) at 2.5 Å resolution and (ii) GlcNAc and ␤-maltose at 1.7Å resolution. The binding studies carried out using surface plasmon resonance indicated that CPGRP-S binds to MDP with a dissociation constant of 10 ؊7 M, whereas the binding affinities Peptidoglycan recognition proteins (PGRPs) 2 of the innate immune system provide the first line of defense to hosts against infecting microbes. These proteins specifically recognize pathogen-associated molecular patterns (PAMPs) such as peptidoglycan (PGN), LPS, lipoteichoic acid (LTA), mycolic acid, and etc. It is well known that PGN is present on the cell walls of both Gram-negative and -positive bacteria. PGN is a polymer of alternating GlcNAc and N-acetylmuramic acid (MurNAc) in a ␤(134) linkage, cross-linked by short peptide stems composed of alternating L-and D-amino acids. The glycan moiety of PGN is conserved among almost all of the bacteria, but the peptide part displays a considerable diversity (1). According to the amino acid residue at number three position of the peptide stem, PGNs are divided into two major categories, (i) lysine type (Lys-type) and (ii) meso-diaminopimelic acid type. The peptidoglycans or their fragments are recognized by the host through innate immunity proteins, particularly PGRPs (2).PGRPs are highly conserved molecules (3-7), which bind to PGN and other PAMPs as well as to their fragments and derivatives with varying affinities (8). There are four types of mammalian PGRPs designated as short PGRP (PGRP-S), PGRP-L, PGRP-I␣, and PGRP-I␤ representing short (molecular mass, 20 -25 kDa), long (molecular mass, up to 90 kDa), and intermediate (molecular mass, 40 -45 kDa), respectively. PGRP-S contains ϳ200 amino acid residues and represents a single PGRP domain, wheraes PGRP-L and PGRP-I also contain a similar C-terminal PGRP domain. PGRP-S shows direct bacterial recognition. The potency of its recognition determines its strength in controlling bacterial infections. Initially, PGRP-S was detected in bone marrow (6), whereas subsequently, it was also observed as a soluble protein in the granules of polymorphonuclear leukocytes (9). This observation implied that PGRP-S might have a role as an antibacterial agent (6). Recently, its presence has also been shown in milk of certain species of mammals (10) and in the intestinal M cells (11). As far as mammary gland secretions are concerned, so far, PGRP-S has been detected in the mammary secretions of porcine and camel only (10). As far as structural studies are concerned, the crystal structures of truncated human PGRP-S (HPGRP-S) (amino acid residues from 9 to 191) and a full-length (amino acid residues from 1 to 171) camel PGRP-S (CPGRP-S) have been reported (12,13) showing truncated HPGRP-S as a monom...

Structural Studies on Molecular Interactions between Camel Peptidoglycan Recognition Protein, CPGRP-S, and Peptidoglycan Moieties N-Acetylglucosamine and N-Acetylmuramic Acid

“…The excellent fittings of both glycan moieties in the subsite with extensive intermolecular contacts indicated that this subsite is formed optimally for interacting with glycan moieties. The previously determined structures of the complexes of CPGRP-S with other glycan moiety-containing PAMPs such as LPS (4), LTA (4), and muramyl dipeptide (5) showed that their glycan moieties also occupied the same pocket as observed in the present structures, whereas other aliphatic attachments of these molecules were adjusted into other parts of the cleft (Fig. 7).…”

“…These steps further reduced the values of R cryst factors to 25.3 and 27.8% for structures GlcNAc and MurNAc, respectively. The Fourier (2 F o Ϫ F c ) and difference Fourier (F o Ϫ F c ) maps computed at this stage showed the presence of significant non-protein electron densities at the ligand binding cleft in molecule C. It may be recalled that the ligand binding site in CPGRP-S is located in molecule C at the C-D contact (4,5). The ligands GlcNAc and MurNAc were fitted well into the electron densities (Fig.…”

“…It is well known that in Gram-negative bacteria, the third residue is meso-diaminopimelic acid, whereas in Gram-positive bacteria, it is L-lysine. To characterize the binding properties of CPGRP-S with PGN and various other PAMPs, we have determined the crystal structures of three complexes of CPGRP-S with lipopolysaccharide (LPS (4)), lipoteichoic acid (LTA (4)), and muramyl dipeptide (MDP (5)). The structures of a few other complexes of different PGRPs such as human PGRP-I␣ (HPGRP-I␣) with muramyl tripeptide (MTP (6)), muramyl pentapeptide (N-acetylmuramyl-alanyl-Glu-Lys-Ala-Ala (7)), and HPGRP-I␤ with muramyl pentapeptide (N-acetyl-D-glucosamine-methyl-2-(acetylamino)-3-O-[(1r)-1-carboxyethyl]-2-deoxy-␤-D-glucopyranoside-Ala-Glu-Lys-Ala-Ala (8)) as well as with Drosophila PGRP-LCa (DPGRP-LCa) and Drosophila PGRP-LCx (DPGRP-LCx) with tracheal cytotoxin (9) have also been reported.…”

Messenger Functions of the Bacterial Cell Wall-derived Muropeptides

“…176-180 Several studies elucidate specific interactions between human PGRPs and peptidoglycan using synthetic peptidoglycan monomers. 178,181-183 …”

Skills in biocatalysis are essential for establishing a sound biotechnological base in India

“…That is an extensive contribution. (Salunke et al, 1985;Banerjee et al, 1994Banerjee et al, , 1996Sankaranarayanan et al, 1996;Singh et al, 2001;Sharma et al, 2011;Rastogi et al, 2014) We need to include many areas like non-aqueous enzymology, enzyme promiscuity and moonlighting proteins in our syllabi and encourage people to work in these areas. The first one offers an opportunity for biocatalyst-based synthesis of industrial and drug intermediates, agrochemicals and biosurfactants (Gupta, 1992;Halling, 1994;Carrea and Riva, 2000;Hudson et al, 2005).…”