Identification of a staphylococcal AgrB segment(s) responsible for group-specific processing of AgrD by gene swapping

The four gene products of the accessory gene regulator (agr) P2 operon of Staphylococcus aureus assemble a quorum-sensing system: AgrA and AgrC resemble a two-component signal transduction system, and AgrB and AgrD are required to produce an autoinducing peptide. Upon activation, this quorum-sensing system positively regulates the transcription of the P2 operon as well as the P3 operon, whose transcript, RNAIII, regulates the expression of virulence genes. Four groups of S. aureus have been identified based on the agr sequences and the group-specific interaction between the autoinducing peptide and AgrC. AgrB is a transmembrane protein involved in the processing of AgrD propeptide, and its interaction with AgrD is also group specific. In this study, a series of chimeric AgrBs were constructed by swapping between group I and group II AgrBs, and these mutants were used to analyze the group-specific segment(s) in AgrB that was responsible for AgrD processing. Our results revealed that the first transmembrane alpha-helix and the extracellular loop 1 of group I AgrB were decisive in the specific processing of group I AgrD. In contrast, two hydrophilic segments of group II AgrB played a crucial role in the group-specific processing of group II AgrD. We also found that several chimeric AgrBs were capable of processing AgrD from both groups, suggesting that all AgrB homologues may utilize the same or a similar mechanism in the processing of AgrDs.