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Analysis of genes encoding two unique type IIa immunoglobulin G-binding proteins expressed by a single group A
M D Boyle1, J Hawlitzky, R Raeder
1Department of Microbiology, Medical College of Ohio, Toledo 43699-0008.
Abstract:
An emm-like gene (emmL) and a fcrA gene from group A streptococcal strain 64/14 (emmL64/14 and fcrA64/14) were amplified by PCR and force cloned into the heat-inducible expression vector pJLA 602. The emmL gene encoded a recombinant protein that bound human IgG1, IgG2, and IgG4 in a nonimmune fashion. This is the reactivity profile of a type IIa IgG-binding protein. The emmL64/14 gene product was antigenically similar to the previously identified high-molecular-weight type IIa IgG-binding protein of strain 64/14 and had an N-terminal sequence identical to that of the wild-type protein. The fcrA gene also encoded a recombinant protein with type IIa functional activity. This protein was similar to the lower-molecular-weight type IIa IgG-binding protein previously isolated from strain 64/14 and was antigenically distinct from the higher-molecular-weight type IIa protein encoded by the emmL64/14 gene. The sequences for both genes including the intervening regions are presented. The emmL gene demonstrates significant homology to other class I emm and emmL genes expressed by opacity factor-negative group A streptococcal isolates. The fcrA gene was found to be homologous to other fcrA genes normally present in opacity factor-positive group A isolates. The sequence upstream of the fcrA gene and the intervening sequence between the end of the fcrA gene and the start of the emmL gene were similar to those reported for other fcrA genes.
Insights
Group A Streptococcus harbors emmL and fcrA genes encoding distinct IgG-binding proteins. These proteins, emmL64/14 and fcrA64/14, exhibit type IIa IgG-binding activity and share homology with known streptococcal genes.
Area of Science:
- Microbiology
- Molecular Biology
- Immunology
Background:
- Group A Streptococcus (GAS) possesses various surface proteins that mediate host immune evasion.
- IgG-binding proteins are crucial virulence factors for GAS, facilitating immune evasion.
- Previous studies identified distinct high- and low-molecular-weight IgG-binding proteins in GAS strain 64/14.
Purpose of the Study:
- To clone and characterize the emm-like (emmL) and fcrA genes from GAS strain 64/14.
- To determine the IgG-binding properties and antigenic similarities of the encoded proteins.
- To analyze the genetic relatedness of emmL64/14 and fcrA64/14 to other streptococcal genes.
Main Methods:
- Polymerase chain reaction (PCR) amplification of emmL and fcrA genes.
- Cloning of amplified genes into a heat-inducible expression vector (pJLA 602).
- Expression and purification of recombinant proteins for functional and antigenic analysis.
Main Results:
- The emmL64/14 gene encoded a recombinant protein binding human IgG subclasses IgG1, IgG2, and IgG4, consistent with type IIa IgG-binding activity.
- The fcrA64/14 gene product also exhibited type IIa functional activity and was antigenically distinct from the emmL64/14 protein.
- emmL64/14 showed homology to class I emm/emmL genes in opacity factor-negative GAS, while fcrA64/14 was homologous to fcrA genes in opacity factor-positive GAS.
Conclusions:
- GAS strain 64/14 possesses at least two distinct genes, emmL and fcrA, encoding type IIa IgG-binding proteins.
- The emmL and fcrA genes likely contribute to GAS virulence through immune evasion mechanisms.
- Comparative genomic analysis reveals distinct evolutionary origins for emmL and fcrA genes in relation to GAS opacity factor production.