PMID:9555900

Citation	Sharma, VK, Hackbarth, CJ, Dickinson, TM and Archer, GL (1998) Interaction of native and mutant MecI repressors with sequences that regulate mecA, the gene encoding penicillin binding protein 2a in methicillin-resistant staphylococci. J. Bacteriol. 180:2160-6
Abstract	Methicillin resistance in staphylococci is mediated by PBP2a, a penicillin binding protein with low affinity for beta-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.
Links	PubMed PMC107144
Keywords	Bacterial Proteins/metabolism; Base Sequence; Carrier Proteins/biosynthesis; Carrier Proteins/genetics; Cloning, Molecular; DNA Primers; Deoxyribonuclease I; Escherichia coli; Gene Expression Regulation, Bacterial; Genes, Bacterial; Hexosyltransferases; Methicillin Resistance/genetics; Molecular Sequence Data; Muramoylpentapeptide Carboxypeptidase/biosynthesis; Muramoylpentapeptide Carboxypeptidase/genetics; Mutagenesis, Site-Directed; Penicillin-Binding Proteins; Peptidyl Transferases; Polymerase Chain Reaction; Promoter Regions, Genetic; Recombinant Proteins/biosynthesis; Recombinant Proteins/metabolism; Repressor Proteins/biosynthesis; Repressor Proteins/genetics; Repressor Proteins/metabolism; Staphylococcus aureus/genetics; Staphylococcus aureus/metabolism; Transcription, Genetic
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Annotations

Gene product	Qualifier	GO Term	Evidence Code	with/from	Aspect	Extension	Notes	Status
STAAU:MECI		GO:0006355: regulation of transcription, DNA-dependent	ECO:0000314:		P		Table 2 shows that in cells transformed with mecI, the beta-galactosidase activity of mecA-lacZ transcriptional fusion can be induced with the presence of CBAP. Furthermore, it is shown in Fig. 2 that mecI binds to a region of DNA between the mecA and mecR1 genes and protects it from DNase.	complete CACAO 4693

References

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