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Immunogenicity and characteristics of M protein released by phage-associated lysin from group-A streptococci types 1
Abstract:
A phage-associated lysin (PAL) was used to release M protein from goup-A streptococci of types 1 and 23. Much of the lysin-released-M protein (LYSIN-M) of both types was of high molecular weight, since LYSIN-M appeared just after the void volume on Sephadex G-200 gel-filtration. Some of the LYSIN-M appeared just after the void volume on Sephadex G-200 gel-filtration. Some of the LYSIN-M of both types was found to be firmly attached to group-A carbohydrate. Type-1 LYSIN-M was partially purified by ammonium-sulfate precipitation followed by absorption and elution from an immunoabsorbent column containing antibody for group-A carbohydrate. Type-23 LYSIN-M was partially purified by precipitation at its isoelectric point, pH 4-9. Rabbits immunised in the footpads with either type-1 or type-23 LYSIN-M responded by producing both precipitins and bactericidal (opsonising) antobodies. Some of the antiesera were absorbed and rendered specific for homologous acid extracts. The LYSIN-M preparations of both types 1 and 23 were originally contaminated with heat-labile antigen(s). Antibodies to these heat-labile antigen(s), which cross-react from type to type, were found in the type-specific antisera distributed by the Center for Disease Control. The specificity of Lancefield typing antisera depends on their being tested with extracts of streptococci prepared at pH 2 and 100 degree C for 10 min. Although LYSIN-M is more difficult to prepare and purify then acid-heat released M protein, it might prove useful for studying the nature of native streptococcal M protein.
Insights
Phage-associated lysin (PAL) releases high-molecular-weight M protein from group A streptococci. This lysin-released M protein (LYSIN-M) can elicit protective antibodies, offering a potential tool for studying native streptococcal M protein.
Area of Science:
- Microbiology
- Immunology
- Protein Chemistry
Background:
- Group A streptococci (GAS) possess M protein, a major virulence factor crucial for bacterial survival and host immune evasion.
- Traditional methods for M protein extraction often involve harsh conditions (acid-heat) that may alter the native protein structure.
- Understanding the native structure of M protein is essential for developing effective vaccines and diagnostics.
Purpose of the Study:
- To investigate the characteristics of M protein released by a phage-associated lysin (PAL).
- To assess the immunogenicity of lysin-released M protein (LYSIN-M) and its potential for generating protective antibodies.
- To compare LYSIN-M with conventionally extracted M protein for studying native streptococcal M protein.
Main Methods:
- Release of M protein from GAS types 1 and 23 using PAL.
- Characterization of LYSIN-M by Sephadex G-200 gel filtration to determine molecular weight.
- Partial purification of LYSIN-M using ammonium sulfate precipitation and immunoabsorbent chromatography (type 1) or isoelectric precipitation (type 23).
- Immunization of rabbits with purified LYSIN-M to generate antisera and assess antibody production (precipitins and bactericidal antibodies).
Main Results:
- LYSIN-M from both types 1 and 23 GAS was predominantly high molecular weight, eluting near the void volume.
- A portion of LYSIN-M was found to be tightly bound to the group A carbohydrate.
- Immunization with LYSIN-M induced both precipitating and bactericidal antibodies in rabbits.
- Antisera showed cross-reactivity, with specific antibodies identified after absorption.
- LYSIN-M preparations contained heat-labile antigens that cross-reacted between types.
Conclusions:
- PAL is effective in releasing high-molecular-weight M protein from GAS, potentially preserving its native conformation.
- LYSIN-M is immunogenic and elicits antibodies capable of opsonization and bactericidal activity.
- While purification is more complex than acid-heat extraction, LYSIN-M offers a valuable alternative for studying native streptococcal M protein structure and function.