Structure, activity and evolution of the group I thiolactone peptide quorum‐sensing system ofStaphylococcus aureus

Abstract

SummaryIn Staphylococcus aureus, the agr locus is responsible for controlling virulence gene expression via quorum sensing. As the blockade of quorum sensing offers a novel strategy for attenuating infection, we sought to gain novel insights into the structure, activity and turnover of the secreted staphylococcal autoinducing peptide (AIP) signal molecules. A series of analogues (including the L-alanine and D-amino acid scanned peptides) was synthesized to determine the functionally critical residues within the S. aureus group I AIP. As a consequence, we established that (i) the group I AIP is inactivated in culture supernatants by the formation of the corresponding methionyl sulphoxide; and (ii) the group I AIP lactam analogue retains the capacity to activate agr, suggesting that covalent modification of the AgrC receptor is not a necessary prerequisite for agr activation. Although each of the D-amino acid scanned AIP analogues retained activity, replacement of the endocyclic amino acid residue (aspartate) located C-terminally to the central cysteine with alanine converted the group I AIP from an activator to a potent inhibitor. The screening of clinical S. aureus isolates for novel AIP groups revealed a variant that differed from the group I AIP by a single amino acid residue (aspartate to tyrosine) in the same position defined as critical by alanine scanning. Although this AIP inhibits group I S. aureus strains, the producer strains possess a functional agr locus dependent on the endogenous peptide and, as such, constitute a fourth S. aureus AIP pheromone group (group IV). The addition of exogenous synthetic AIPs to S. aureus inhibited the production of toxic shock syndrome toxin (TSST-1) and enterotoxin C3, confirming the potential of quorum-sensing blockade as a therapeutic strategy.

Inactivation of a bacterial virulence pheromone by phagocyte-derived oxidants: New role for the NADPH oxidase in host defense

“…3C), and the biologic significance of this modification in the absence of oxidation is presently unknown. Even though the methionine is not essential for the function of this AIP (33), our data confirm the loss of biologic activity observed with the methionine sulfoxide form of this peptide purified from culture supernatants (34). This finding suggests that oxidation may alter either the conformation or polarity of the peptide so its interaction with the bacterial receptor agrC is impaired.…”

Exploiting Quorum Sensing To Confuse Bacterial Pathogens

“…The tail of the AIP molecule is required only for activation, as mutation of the tail amino acid residues or removal of the tail did not alter inhibition of the agr response in heterologous groups despite eliminating self-activation activity (130,219). Numerous SAR studies have mapped amino acid residues critical for AIP activity, and overall the results demonstrate that critical amino acids differ between AIP groups (218)(219)(220). For example, in AIP-I the endocyclic aspartic acid residue adjacent to the cysteine residue is critical, as replacement of this amino acid with alanine converts AIP-I to a potent inhibitor of AgrC of groups I and IV (218,219).…”

“…The molecular determinants for AIP specificity have been shown to reside entirely in the interaction between the AIP signal and the AgrC receptor (130). Both cognate activation and heterologous inhibition activities require the ring structure of AIP, as linear AIPs have no detectable activity in any agr group (219,220), but the requirement of the thioester linkage has been debated (218)(219)(220). The tail of the AIP molecule is required only for activation, as mutation of the tail amino acid residues or removal of the tail did not alter inhibition of the agr response in heterologous groups despite eliminating self-activation activity (130,219).…”

“…This may help to explain why residues critical for activity differ between AIP groups. Systematic mutation of native AIP molecules has generated analogues with various changes in activity, including reduced activation activity, enhanced activation activity, loss of all activity, conversion of self-activation to self-inhibition activity, and a unique case of conversion of cross-inhibition to cross-activation activity (218)(219)(220)(221). Combining multiple mutations known to provide antagonist activity, for example, truncating the tail of a selfinhibitory AIP-I mutant, has also been shown to have potential in the generation of stronger inhibitors that function against all agr groups (218).…”

agr receptor mutants reveal distinct modes of inhibition by staphylococcal autoinducing peptides

“…The agr quorum-sensing system allows S. aureus to navigate its host and strategically switch, in response to bacterial density, between attachment and aggressive lifestyles during the course of infection. The agr autoinducing peptide (AIP) is 7-9 residues in length and contains a 5-membered ring in which the C terminus forms a thiolactone bond with a conserved, central cysteine, a structure that is critical for activity (2)(3)(4). Binding of the AIP, via a conserved C-terminal hydrophobic patch (5) and additional specific contacts (6,7), to the hexahelical transmembrane (TM) sensor domain of its cognate receptor, AgrC, results in trans-autophosphorylation of the receptor via its cytoplasmic histidine kinase (HK) domain.…”