Interaction of Native and Mutant MecI Repressors with Sequences That Regulate mecA, the Gene Encoding Penicillin Binding Protein 2a in Methicillin-Resistant Staphylococci
AUTOR(ES)
Sharma, Vijay K.
FONTE
American Society for Microbiology
RESUMO
Methicillin resistance in staphylococci is mediated by PBP2a, a penicillin binding protein with low affinity for β-lactam antibiotics. The gene encoding PBP2a, mecA, is transcriptionally regulated in some clinical isolates by mecR1 and mecI, genes divergently transcribed from mecA that encode a signal transducer and repressor, respectively. The biochemical basis of MecI-mediated mecA transcriptional repression was investigated by using purified MecI. In DNase I protection studies, MecI protected a 30-bp palindrome encompassing the predicted mecA −10 and the mecR1 −35 promoter sequences. The larger palindrome contained 15 bp of dyad symmetry within which was a smaller 6-bp palindrome. Electrophoretic mobility shift assays established a requirement for the entire 15-bp half-site for initial repressor binding. Fragments containing the 30-bp palindrome and the entire mecA-mecR1 intergenic region were retarded in gels as multiple discrete bands varying in molecular size, characteristic of cooperative DNA binding. Glutaraldehyde cross-linking confirmed oligomerization of repressor in solution. A naturally occurring MecI mutant (MecI*; D39G) repressed mecA transcription sixfold less well than the wild type in vivo. Although MecI* protected the same target sequences and exhibited similar gel shift patterns to MecI, 5- to 10-fold more protein was required. MecI* exhibited defective oligomerization in solution, suggesting that the MecI amino terminus is important in protein-protein interactions and that protein oligomerization is necessary for optimum repression.
ACESSO AO ARTIGO
http://www.pubmedcentral.nih.gov/articlerender.fcgi?artid=107144Documentos Relacionados
- Survey of the methicillin resistance-associated genes mecA, mecR1-mecI, and femA-femB in clinical isolates of methicillin-resistant Staphylococcus aureus.
- Analysis of Diversity of Mutations in the mecI Gene and mecA Promoter/Operator Region of Methicillin-Resistant Staphylococcus aureus and Staphylococcus epidermidis
- Survey of methicillin-resistant clinical strains of coagulase-negative staphylococci for mecA gene distribution.
- Expression and inducibility in Staphylococcus aureus of the mecA gene, which encodes a methicillin-resistant S. aureus-specific penicillin-binding protein.
- Rapid detection of the mecA gene in methicillin-resistant staphylococci by enzymatic detection of polymerase chain reaction products.