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Journal of Bacteriology, September 2005, p. 5955-5966, Vol. 187, No. 17
0021-9193/05/$08.00+0     doi:10.1128/JB.187.17.5955-5966.2005
Copyright © 2005, American Society for Microbiology. All Rights Reserved.

Identification of Residues Responsible for the Defective Virulence Gene Regulator Mga Produced by a Natural Mutant of Streptococcus pyogenes

Cheryl M. Vahling and Kevin S. McIver^*

Department of Microbiology, University of Texas Southwestern Medical Center, Dallas, Texas 75390

Received 8 April 2005/ Accepted 14 June 2005

Mga is a transcriptional regulator in the pathogen Streptococcus pyogenes that positively activates several important virulence genes involved in colonization and immune evasion in the human host. A naturally occurring mutant of Mga that is defective in its ability to activate transcription has been identified in the serotype M50 strain B514-Sm. Sequence alignment of the defective M50 Mga with the fully functional Mga from serotypes M4 and M49 revealed only three amino acid changes that might result in a defective protein. Electrophoretic mobility shift assays using purified M50 and M4 maltose binding protein-Mga found that both exhibited DNA-binding activity towards regulated promoters. Thus, the significance of each residue for the functionality of M50 Mga was explored through introduction of "gain-of-function" mutations based on M4 Mga. Transcriptional studies of the mutant alleles under both constitutive (PrpsL) and autoactivated (Pmga4) promoters illustrated that an arginine-to-methionine change at position 461 of M50 Mga protein fully restored activation of downstream genes. Western blot analyses of steady-state Mga levels suggest that the M461 residue may play a role in overall conformation and protein stability of Mga. However, despite the conservation of the M461 protein among all other Mga proteins, it does not appear to be necessary for activity in a divergent M6 Mga. These studies highlight the potential differences that exist between divergent Mga proteins in this important human pathogen.

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* Corresponding author. Mailing address: Department of Microbiology, UT Southwestern Medical Center, 5323 Harry Hines Blvd., Dallas, TX 75390-9048. Phone: (214) 648-1255. Fax: (214) 648-5907. E-mail: kevin.mciver{at}utsouthwestern.edu.

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Journal of Bacteriology, September 2005, p. 5955-5966, Vol. 187, No. 17
0021-9193/05/$08.00+0     doi:10.1128/JB.187.17.5955-5966.2005
Copyright © 2005, American Society for Microbiology. All Rights Reserved.

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