Studies of the repressor (BlaI) of β‐lactamase synthesis in <i>Staphylococcus aureus</i>

Gregory, Philip D.; Lewis, Richard A.; Curnock, S.P.; Dyke, K. G. H.

doi:10.1046/j.1365-2958.1997.4051770.x

Cited by 87 publications

(109 citation statements)

References 28 publications

(31 reference statements)

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“…This event triggers the activation of the putative cytoplasmic metalloprotease motif of BlaR1 by self-proteolysis (8,12). In S. aureus, the ␤-lactamase induction correlates with BlaI proteolysis between residues Asn 101 and Phe 102 , and it is postulated that the activated form of BlaR1 proteolyses BlaI (12)(13)(14)(15). The blaR2 gene could be involved in this process or in the activation of BlaR1, but our recent studies in a recombinant Bacillus subtilis harboring a plasmid carrying the bla divergeon show contrasting results with those in S. aureus and suggest that the inactivation of BlaI in Bacillus is mediated by the binding of a coactivator (16).…”

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confidence: 99%

Dimerization and DNA Binding Properties of theBacillus licheniformis 749/I BlaI Repressor

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Vreuls

Herman

et al. 2003

Journal of Biological Chemistry

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In the absence of penicillin, the ␤-lactamase encoding gene blaP of Bacillus licheniformis 749/I is negatively regulated by the transcriptional repressor BlaI. Three palindromic operator regions are recognized by BlaI: two in the blaP promoter (OP1 and OP2) and one (OP3) in the promoter of the blaI-blaR1 operon. In this study, the dissociation constant of the purified BlaI dimer was estimated at 25 M by equilibrium ultracentrifugation. Quantitative Western blot analysis indicates that the intracellular concentrations of BlaI in B. licheniformis 749/I and Bacillus subtilis transformed by a multicopy plasmid harboring the ␤-lactamase locus (blaP-blaIblaR1) were lower than (1.9 M) or in the same range as (75 M) the dissociation constant, respectively. This suggests that BlaI is partially dimeric in the cytoplasm of these strains and interacts in vivo with its operators as a preformed dimer. This hypothesis is supported by band shift assays on an operator containing a randomized half-operator sequence. The global dissociation constants of the operator-BlaI dimer complexes were measured by band shift assays and estimated as K dOP1 ‫؍‬ 1.7 ؎ 0.5 10 The blaP gene encodes the class A ␤-lactamase of Bacillus licheniformis 749/I. In the absence of ␤-lactams antibiotics, the BlaI repressor prevents the transcription of the blaP gene (1-3). Two additional genes, blaR1 and blaR2, are also involved in the induction of the ␤-lactamase synthesis (4, 5). The blaP, blaI, and blaR1 genes are clustered in a divergeon (bla divergeon) in which blaI and blaR1 form an operon (Fig. 1). The blaR1 gene encodes a transmembrane protein that acts as penicillin receptor (6 -8), and blaR2 is not identified yet and is not linked to the bla divergeon. The BlaR2 protein is essential for the inactivation of BlaI. In Staphylococcus aureus, the blaZ and mecA genes, encoding respectively a ␤-lactamase and the low affinity penicillinbinding protein 2Ј, are regulated by similar elements (9, 10).The blaI gene encodes a 128-residue protein characterized by two functional and separate domains (11). The DNA-binding domain, a helix-turn-helix recognition motif, is located in the N-terminal region, and the dimerization domain is in the Cterminal region. After deletion of the C-terminal domain, the repressor becomes unable to dimerize and to form a stable complex with its operators. DNase footprinting experiments and filter binding reactions revealed the presence of three regulatory regions that are recognized specifically by BlaI (11). These operators present a 23-bp-long dyad symmetry and show the deduced 5Ј-AAAGTATTACATATGTAACNTTT-3Ј consensus sequence (Fig. 1).In the presence of ␤-lactam antibiotics, the C-terminal domain of BlaR1 is acylated (6, 7). This event triggers the activation of the putative cytoplasmic metalloprotease motif of BlaR1 by self-proteolysis (8, 12). In S. aureus, the ␤-lactamase induction correlates with BlaI proteolysis between residues Asn 101 and Phe 102 , and it is postulated that the activated form of BlaR1 proteolyses BlaI (12-...

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confidence: 99%

Dimerization and DNA Binding Properties of theBacillus licheniformis 749/I BlaI Repressor

Filée

Vreuls

Herman

et al. 2003

Journal of Biological Chemistry

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“…Recent studies have demonstrated that the signaling pathway responsible for β-lactamase synthesis requires sequential cleavage of the regulatory proteins BlaR1 and BlaI. Following exposure to β-lactams, BlaR1, a transmembrane sensor-transducer, cleaves itself (17,18). Zhang et al (18) hypothesize that the cleaved protein functions as a protease that cleaves the repressor BlaI, directly or indirectly (an additional protein, BlaR2, may be involved in this pathway) and allows blaZ to synthesize enzyme.…”

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confidence: 99%

Antimicrobial resistance: the example of Staphylococcus aureus

Lowy¹

2003

J. Clin. Invest.

394

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“…CopY, a member of the Bla/ Mec repressor family, is the archetypical prokaryotic copper regulated repressor [20]. In the DNA binding state, CopY binds one Zn(II) ion in a -CxCxxxxCxC-motif.…”

Section: Regulation Of the Cop Operon And Copymentioning

confidence: 99%

“…To maintain DNA binding activity Zn(II) must be bound to the protein [21]. Within the Bla/Mec family of repressors the affinity for DNA is strongly linked to the formation of dimers [20]. Cu(I) ions displace the Zn(II) in the CopY metal binding site, prompting the protein to lose affinity for DNA [16].…”

Section: The Structure Of Copymentioning

confidence: 99%