Peptidoglycan and lipopolysaccharide bind to the same binding site on lymphocytes (original) (raw)
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Journal of Biological Chemistry, 1991
One dominant binding site (70 kDa 6.5 PI protein) for bacterial cell wall peptidoglycan (PGN), a macrophage activator and polyclonal B cell mitogen, was demonstrated on mouse B and T lymphocytes and macrophages by photoaffinity cross-linking and two-dimensional polyacrylamide gel electrophoresis. This binding site was not present on erythrocytes. The binding was specific for polymeric PGN and was competitively inhibited by unlabeled PGN with ICs0 = 48 pg/ ml (0.38 p~). The binding was partially inhibited by 0-acetylated PGN monomers (IC60 = 469 pg/ml, 521 NM), dextran sulfate (ICso = 1024 pg/ml, 124 p~) , and (GlcNAc)3 (ICso = 6.6 mg/ml, 10 mM), and was not inhibited by non-0-acetylated PGN monomers and dimers, muramyl dipeptide, PGN pentapeptide, GlcNAc, teichoic acid, protein A, and gelatin. The cell surface location of the 70-kDa PGN-binding protein was indicated by the ability of PGN to bind to this protein in intact metabolically inactive cells (at 4 "C and in the presence of 0.1% NaN3) and by the ability to extract the 70-kDa PGN-binding protein from viable B lymphocytes by noncytotoxic concentration of n-octyl-8-D-ghcopyranoside.
Journal of Biological Chemistry, 2003
The C-terminal region (cA) of the major autolysin AcmA of Lactococcus lactis contains three highly similar repeated regions of 45 amino acid residues (LysM domains), which are separated by nonhomologous sequences. The cA domain could be deleted without destroying the cell wall-hydrolyzing activity of the enzyme in vitro. This AcmA derivative was capable neither of binding to lactococcal cells nor of lysing these cells while separation of the producer cells was incomplete. The cA domain and a chimeric protein consisting of cA fused to the C terminus of MSA2, a malaria parasite surface antigen, bound to lactococcal cells specifically via cA. The fusion protein also bound to many other Gram-positive bacteria. By chemical treatment of purified cell walls of L. lactis and Bacillus subtilis, peptidoglycan was identified as the cell wall component interacting with cA. Immunofluorescence studies showed that binding is on specific locations on the surface of L. lactis, Enterococcus faecalis, Streptococcus thermophilus, B. subtilis, Lactobacillus sake, and Lactobacillus casei cells. Based on these studies, we propose that LysM-type repeats bind to peptidoglycan and that binding is hindered by other cell wall constituents, resulting in localized binding of AcmA. Lipoteichoic acid is a candidate hindering component. For L. lactis SK110, it is shown that lipoteichoic acids are not uniformly distributed over the cell surface and are mainly present at sites where no MSA2cA binding is observed.
The Journal of Infectious Diseases, 2001
Infection with gram-positive bacteria is a major cause of pneumonia. Surfactant proteins A (SP-A) and D (SP-D) are thought to play an important role in the innate immunity of the lung. Both proteins can bind to gram-positive bacteria. Until now, it was not known with which surface component(s) of gram-positive bacteria SPA and SP-D interact. Lipoteichoic acid (LTA) and peptidoglycan (PepG) are components of the cell wall of gram-positive bacteria. By use of a solid phase-based binding assay, LTA of Bacillus subtilis was shown to be bound by SP-D but not by SPA. Unmodified PepG of Staphylococcus aureus was bound by SP-D. SP-D binding to both LTA and PepG was calcium dependent and carbohydrate inhibitable. These results indicate that SP-D interacts with gram-positive bacteria via binding to the cell wall components LTA and PepG and that the carbohydrate recognition domain is responsible for this binding. It is well known that gram-positive bacteria, like Staphylococcus aureus and especially Streptococcus pneumoniae, can cause pulmonary inflammation [1, 2]. An evaluation of adult respiratory distress syndrome cases found that more than one-third were caused by infection with gram-positive bacteria [3]. Furthermore, infection with pneumococci, for example, is a leading cause of death, with a mortality rate of 40% in otherwise healthy elderly individuals [4]. Lipoteichoic acid (LTA) and peptidoglycan (PepG) are 2 major cell wall components of gram-positive organisms [5] and can induce an inflammatory response. Besides pulmonary infection, gram-positive bacteria can initiate septic shock, which is induced by LTA and PepG [6]. PepG is an alternating blinked (1, 4) N-acetylmuramyl and N-acetylglucosaminyl glycan whose residues are cross-linked via short peptides. Gram-positive bacteria are encased by multiple layers of cross-linked PepG. It is, therefore, the most prevalent component of the cell wall of gram-positive bacteria, accounting for up to 40% of the weight of the bacterial cell wall [7]. LTAs are members of a group of structurally related macroamphiphiles. They are glycolipids composed of a hydrophobic diacylglycerol membrane anchor and a hydrophilic group pointing to the bacterial surface. This hydrophilic side chain varies among different bacterial species. LTA is present in most
PLoS ONE, 2013
Peptidoglycan recognition proteins (PGRPs) are part of the innate immune system. The 19 kDa Short PGRP (PGRP-S) is one of the four mammalian PGRPs. The concentration of PGRP-S in camel (CPGRP-S) has been shown to increase considerably during mastitis. The structure of CPGRP-S consists of four protein molecules designated as A, B, C and D forming stable intermolecular contacts, A-B and C-D. The A-B and C-D interfaces are located on the opposite sides of the same monomer leading to the the formation of a linear chain with alternating A-B and C-D contacts. Two ligand binding sites, one at C-D contact and another at A-B contact have been observed. CPGRP-S binds to the components of bacterial cell wall molecules such as lipopolysaccharide (LPS), lipoteichoic acid (LTA), and peptidoglycan (PGN) from both Gram-positive and Gramnegative bacteria. It also binds to fatty acids including mycolic acid of the Mycobacterium tuberculosis (Mtb). Previous structural studies of binary complexes of CPGRP-S with LPS and stearic acid (SA) have shown that LPS binds to CPGRP-S at C-D contact (Site-1) while SA binds to it at the A-B contact (Site-2). The binding studies using surface plasmon resonance showed that LPS and SA bound to CPGRP-S in the presence of each other. The structure determination of the ternary complex showed that LPS and SA bound to CPGRP-S at Site-1 and Site-2 respectively. LPS formed 13 hydrogen bonds and 159 van der Waals contacts (distances #4.2 Å) while SA formed 56 van der Waals contacts. The ELISA test showed that increased levels of productions of pro-inflammatory cytokines TNF-a and IFN-c due to LPS and SA decreased considerably upon the addition of CPGRP-S.
Binding of Bacterial Peptidoglycan to CD14
Journal of Biological Chemistry, 1998
The hypothesis that soluble peptidoglycan (sPGN, a macrophage-activator from Gram-positive bacteria) binds to CD14 (a lipopolysaccharide (LPS) receptor) was tested. sPGN specifically bound to CD14 in the following three assays: binding of soluble 32 P-CD14 (sCD14) to agarose-immobilized sPGN, enzyme-linked immunosorbent assay, and photoaffinity cross-linking. sCD14 also specifically bound to agarose-immobilized muramyl dipeptide or GlcNAc-muramyl dipeptide but not to PGN pentapeptide. Binding of sCD14 to both sPGN and ReLPS (where ReLPS is LPS from Salmonella minnesota Re 595) was competitively inhibited by unlabeled sCD14, 1-152 N-terminal fragment of sCD14, sPGN, smooth LPS, ReLPS, lipid A, and lipoteichoic acid but not by dextran, dextran sulfate, heparin, ribitol teichoic acid, or soluble low molecular weight PGN fragments. Binding of sCD14 to sPGN was slower than to ReLPS but of higher affinity (K D ؍ 25 nM versus 41 nM). LPS-binding protein (LBP) increased the binding of sCD14 to sPGN by adding another lower affinity K D and another higher B max , but for ReLPS, LBP increased the affinity of binding by yielding two K D with significantly higher affinity (7.1 and 27 nM). LBP also enhanced inhibition of sCD14 binding by LPS, ReLPS, and lipid A. Binding of sCD14 to both sPGN and ReLPS was inhibited by anti-CD14 MEM-18 mAb, but other anti-CD14 mAbs showed differential inhibition, suggesting conformational binding sites on CD14 for sPGN and LPS, that are partially identical and partially different. Peptidoglycan (PGN) 1 is a polymer of alternating GlcNAc and MurNAc cross-linked by short peptides, present in the cell walls of all bacteria (1). PGN, similar to lipopolysaccharide (LPS) from the cell walls of Gram-negative bacteria, can reproduce most of the clinical manifestations of bacterial infections, including fever, acute-phase response,
Lipopolysaccharide (LPS)-binding Proteins BPI and LBP Form Different Types of Complexes with LPS
Journal of Biological Chemistry, 1997
Lipopolysaccharide (LPS)-binding protein (LBP) and bactericidal/permeability-increasing protein (BPI) are closely related LPS-binding proteins whose binding to LPS has markedly different functional consequences. To gain better insight into the possible basis of these functional differences, the physical properties of LBP-LPS and BPI-LPS complexes have been compared in this study by sedimentation, light scattering, and fluorescence analyses. These studies reveal dramatic differences in the physical properties of LPS complexed to LBP versus BPI. They suggest that of the two proteins, only LBP can disperse LPS aggegates. However, BPI can enhance both the sedimentation velocity and apparent size of LPS aggregates while inhibiting LPS-LBP binding even at very low (1:40 to 1:20) BPI:LPS molar ratios. The lipopolysaccharide (LPS)-binding protein 1 (LBP) and the bactericidal/permeability-increasing protein (BPI) are both LPS-interactive mammalian proteins with approximately 45% amino acid sequence identity (1, 2). LPS is considered to be the principal component of Gram-negative bacteria that alerts the host to invading bacteria and triggers defensive responses (3, 4). These responses are usually beneficial and effective but may also become excessive and lead to endotoxic shock (3-5). Both LBP and BPI modulate the bioactivity of LPS (2, 3, 5). LBP is a plasma protein that catalyzes the transfer of LPS from LPS aggregates to other LPS-binding proteins (3, 6-9). Prominent among these is CD14, a surface molecule of myeloid cells that is also present in the circulation as a soluble protein. LBP and CD14 together represent the main pathway by which cells recognize low concentrations of LPS and are stimulated to respond to Gram-negative bacteria (3, 10, 11). In contrast to the LPS-stimulatory properties of LBP, binding of LPS by BPI results in inhibition of the bioactivities of LPS (2). BPI is produced by polymorphonuclear leukocytes and stored in its azurophilic granules (12, 13). It contributes substantially to both the intracellular and extracellular antibacterial activity of polymorphonuclear leukocyte-rich inflammatory exudates toward Gram-negative bacteria (14, 15). The high affinity of BPI
Purification and characterization of murine lipopolysaccharide-binding protein
Infection and Immunity, 1993
The serum protein lipopolysaccharide (LPS)-binding protein (LBP) seems to play an important role in regulating host responses to LPS. Complexes of LPS and LBP form in serum and stimulate monocytes, macrophages, or polymorphonuclear leukocytes after binding to CD14. Previous reports have described the structure and properties of LBP from human and rabbit sera. Since mice are used in some experimental models of endotoxemia or gram-negative bacterial infections, information is needed about the properties of murine LBP. Murine LBP was purified by ion-exchange chromatography and high-pressure liquid chromatography; its NH2-terminal sequence (TNPGLVTRIT) was very similar to those of human and rabbit LBPs (80 to 90% amino acid identity). Murine LBP resembled LBPs from other species in that it promoted the binding of LPS to monocytes and enhanced the sensitivity of monocytes to LPS at least 100-fold. Mouse LBP, like rabbit and human LBPs, was found to be an acute-phase protein. Further in v...
Molecular Immunology, 1990
Tritium-labeled lipopolysaccharide interacted specifically and reversibly with mouse peritoneal macrophages. The binding was higher at 22'C than at 4"C, but was no longer observable at 37'C. The specificity of the interaction (inhibition with unlabeled LPS) was strictly dependent on the presence of serum, and required divalent cations. The binding was saturable. The specific binding sites of peritoneal macrophages were saturated with 6-9 x IO6 LPS molecules/cell, and those of macrophage-like cell lines with 2-3 x 10" molecules/cell.