During sequencing of an 11.5 kb genomic region of a serotype M49 group A streptococcal (GAS) strain, a series of genes were identified including nra (negative regulator of GAS). Transcriptional analysis of the region revealed that nra was primarily monocistronically transcribed. Polycistronic expression was found for the three open reading frames (ORFs) downstream and for the four ORFs upstream of nra. The deduced Nra protein sequence exhibited 62% homology to the GAS RofA positive regulator. In contrast to RofA, Nra was found to be a negative regulator of its own expression and that of the two adjacent operons by analysis of insertional inactivation mutants. By polymerase chain reaction and hybridization assays of 10 different GAS serotypes, the genomic presence of nra, rofA or both was demonstrated. Nra-regulated genes include the fibronectin-binding protein F2 gene (prtF2) and a novel collagen-binding protein (cpa). The Cpa polypeptide was purified as a recombinant maltose-binding protein fusion and shown to bind type I collagen but not fibronectin. In accordance with nra acting as a negative regulator of prtF2 and cpa, levels of attachment of the nra mutant strain to immobilized collagen and fibronectin was increased above wild-type levels. In addition, nra was also found to regulate negatively (four- to 16-fold) the global positive regulator gene, mga. Using a strain carrying a chromosomally integrated duplication of the nra 3′ end and an nra–luciferase reporter gene transcriptional fusion, nra expression was observed to reach its maximum during late logarithmic growth phase, while no significant influence of atmospheric conditions could be distinguished clearly.
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Document Type: Research Article
Department of Medical Microbiology and Hygiene, University Hospital Ulm, Robert-Koch-Str. 8, D-89081 Ulm, Germany.,
Institute of Medical Microbiology, University of Jena, Semmelweisstr. 4, D-07740 Jena, Germany.
February 1, 1999