Relation between wall teichoic acid content and Concanavalin A binding in Bacillus subtilis 168
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It has been known for some time that Concanavalin A will agglutinate bacteria whose wall teichoic acids contain a-linked glucosyl substituents [1]. The ability of the lectin to precipitate appropriately substituted teichoic acids has been used in structural [2] and preparative [3] studies and its interaction with whole bacteria or isolated walls has been studied in attempts to gain information on the location of teichoic acids in the cell wall and at the bacterial surface [4,5]. Interpretation of the latter studies has been complicated by the lack of any information on how the amount of Concanavalin A bound is related to the amount of teichoic acid present in the bacteria, or whether variations in the amount of teichoic acid present affect both the amount bound and the affinity of binding. We have recently [6] shown that the amount of teichoic acid present in walls of B. subtilis 168 can be controlled by growing the bacteria under balanced conditions with varied phosphate supply. We now report a study of the interaction of these bacteria with Concanavalin A. Our results show that the association constant of the binding reaction is little affected by variation in teichoic acid content but the amount of Concanavalin A bound increases in proportion to the amount of teichoic acid present although it ap-Keywords:
Teichoic acid
It has been known for some time that Concanavalin A will agglutinate bacteria whose wall teichoic acids contain a-linked glucosyl substituents [1]. The ability of the lectin to precipitate appropriately substituted teichoic acids has been used in structural [2] and preparative [3] studies and its interaction with whole bacteria or isolated walls has been studied in attempts to gain information on the location of teichoic acids in the cell wall and at the bacterial surface [4,5]. Interpretation of the latter studies has been complicated by the lack of any information on how the amount of Concanavalin A bound is related to the amount of teichoic acid present in the bacteria, or whether variations in the amount of teichoic acid present affect both the amount bound and the affinity of binding. We have recently [6] shown that the amount of teichoic acid present in walls of B. subtilis 168 can be controlled by growing the bacteria under balanced conditions with varied phosphate supply. We now report a study of the interaction of these bacteria with Concanavalin A. Our results show that the association constant of the binding reaction is little affected by variation in teichoic acid content but the amount of Concanavalin A bound increases in proportion to the amount of teichoic acid present although it ap-
Teichoic acid
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Staphylococcus aureus is resistant to α-defensins, antimicrobial peptides that play an important role in oxygen-independent killing of human neutrophils. The dlt operon mediates d-alanine incorporation into teichoic acids in the staphylococcal cell envelope and is a determinant of defensin resistance. By using S. aureus wild-type (WT) and Dlt− bacteria, the relative contributions of oxygen-dependent and -independent antimicrobial phagocyte components were analyzed. The Dlt− strain was efficiently killed by human neutrophils even in the absence of a functional respiratory burst, whereas the killing of the WT organism was strongly diminished when the respiratory burst was inhibited. Human monocytes, which do not produce defensins, inactivated the WT and Dlt− bacteria with similar efficiencies. In addition, mice injected with the Dlt− strain had significantly lower rates of sepsis and septic arthritis and fewer bacteria in the kidneys, compared with mice infected with the WT strain
Teichoic acid
Respiratory burst
Phagocyte
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Human sera were found to contain antibodies precipitating with each of two samples of teichoic acid of Staphylococcus aureus prior to immunization; these antibodies were probably formed as a result of contact or infection with this microorganism. Injection of teichoic acid into two individuals resulted in a rise in circulating antibody to teichoic acid; a third subject probably had a primary response to alpha-teichoic acid. Quantitative precipitin and agar diffusion studies revealed the presence of two distinct antibodies in the sera and showed that each specimen of teichoic acid was a mixture of two polymers an alpha-linked N-acetylglucosaminyl-ribitol polymer and a beta-linked N-acetylglucosaminyl-ribitol polymer, termed alpha- and beta-teichoic acids respectively. The alpha-teichoic acid anti-alpha-teichoic acid system was inhibited best by alpha-linked glucosaminides and the beta-anti-beta-teichoic acid system was inhibited best by a beta-linked glucosaminide. The alpha- and (beta-teichoic acids could be separated from each other by specific precipitation under appropriate conditions and recovered from the washed specific precipitates. The existence of two distinct teichoic acid polymers raises important questions as to cell wall structure and the biosynthesis of the teichoic acids.
Teichoic acid
Precipitin
Lipoteichoic acid
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The interaction of Concanavalin A with teichoic acids from S. aureus and S. epieermidis has been studied by gel diffusion and precipitation in a fluid system. It was found that Concanavalin A reacted with alpha-glucosylated but not with beta-glucosylated teichoic acids from S. epidermidis . It was similarly found that Concanavalin A reacted only with the alpha-linked N-acetylglucosamine teichoic acids from strains of S. aureus . These observations were substantiated by quantitative precipitin and hapten inhibition studies using specific antisera to the staphylococcal strains studied. The usefulness of the interaction of Concanavalin A with teichoic acids in studying the immunochemistry of these antigens was demonstrated.
Teichoic acid
Hapten
Precipitin
Immunochemistry
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Membranes from Bacillus subtilis W23 synthesized a lipid precursor of the linkage unit that attaches teichoic acid to the cell wall. It contained glycerophosphoryl-N-acetylglucosamine, linked through an acid-labile bond to a lipid.
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Linkage (software)
Lipid II
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Teichoic acid
Bacillus (shape)
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Abstract The interaction of Concanavalin A with teichoic acids from S. aureus and S. epieermidis has been studied by gel diffusion and precipitation in a fluid system. It was found that Concanavalin A reacted with alpha-glucosylated but not with beta-glucosylated teichoic acids from S. epidermidis. It was similarly found that Concanavalin A reacted only with the alpha-linked N-acetylglucosamine teichoic acids from strains of S. aureus. These observations were substantiated by quantitative precipitin and hapten inhibition studies using specific antisera to the staphylococcal strains studied. The usefulness of the interaction of Concanavalin A with teichoic acids in studying the immunochemistry of these antigens was demonstrated.
Teichoic acid
Precipitin
Hapten
Immunochemistry
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Interactions between concanavalin A and cell wall digests of Bacillus subtilis 168 resulted in insoluble complexes as observed by double gel diffusion, turbidity, and analysis of the precipitate. The macromolecular constituent of the cell walls complexing with concanavalin A was the polyglucosylglycerol phosphate teichoic acid. The complex exhibited two p H optima: 3.1 and 7.4. The complex could be dissociated by saccharides which bind to concanavalin A. In contrast to concanavalin A-neutral polysaccharide complexes, formation of the concanavalin A-wall complex was inhibited by salts. It was subsequently shown that salts induce conformational changes in cell wall digests. The data suggested that for complex formation to occur a rigid rod conformation in the glucosylated teichoic acid is probably necessary. Concanavalin A can be used as a probe to study structural features of bacterial cell walls.
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Teichoic acid
Trichloroacetic acid
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Hydrolysis of the cell wall of Bacillus subtilis 168 by autolysins or lysozyme resulted in the exposure of glucosylated teichoic acid molecules as evidenced by increased precipitation of [14C] concanavalin A. The number of concanavalin A-reactive sites increased significantly after only limited enzymatic digestion of the walls. Quantitative analyses of [14C] concanavalin A-treated wall or wall hydrolysate complexes indicate that approximately one-half of the teichoic acid molecules are surface-exposed, whereas the remainder are probably embedded within the peptidoglycan matrix. Treatment of the cell walls with sodium dodecyl sulfate or Triton X-100 did not result in new concanavalin A-reactive sites. Partial autolysis diminished the ability of the cell walls to adsorb bacteriophage phi25. Fluorescein-labeled concanavalin A bound intensely over the entire surface of growing B. subtilis 168 cells, suggesting that teichoic acid molecules are located on the total solvent-exposed surface area of the bacteria.
Teichoic acid
Autolysis (biology)
Autolysin
Cell envelope
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