title: Identification of ligand specificity determinants in AgrC, the Staphylococcus aureus quorum-sensing receptor.
abstract: Activation of the agr system, a major regulator of staphylococcal virulence, is initiated by the binding of a specific autoinducing peptide (AIP) to the extracellular domain of AgrC, a classical receptor histidine protein kinase. There are four known agr specificity groups in Staphylococcus aureus, and we have previously localized the determinant of AIP receptor specificity to the C-terminal half of the AgrC sensor domain. We have now identified the specific amino acid residues that determine ligand activation specificity for agr groups I and IV, the two most closely related. Comparison of the AgrC-I and AgrC-IV sequences revealed a set of five divergent residues in the region of the second extracellular loop of the receptor that could be responsible. Accordingly, we exchanged these residues between AgrC-I and AgrC-IV and tested the resulting constructs for activation by the respective AIPs, measuring activation kinetics with a transcriptional fusion of blaZ to the principal agr promoter, P3. Exchange of all five residues caused a complete switch in receptor specificity. Replacement of two of the AgrC-IV residues by the corresponding residues in AgrC-I caused the receptor to be activated by AIP-I nearly as well as the wild type AgrC-I receptor. Replacement of two different AgrC-I residues by the corresponding AgrC-IV residues broadened receptor recognition specificity to include both AIPs. Various types of intermediate activity were observed with other replacement mutations. Preliminary characterization of the AgrC-I-AIP-I interaction suggests that ligand specificity may be sterically determined.
authors.affiliation: Molecular Pathogenesis Program and Departments of Microbiology and Medicine, the Kimmel Center for Biology and Medicine of the Skirball Institute, New York University School of Medicine, New York, NY 10016, USA.
authors.forename: Edward
authors.initials: E
authors.lastname: Geisinger
authors.forename: Elizabeth A
authors.initials: EA
authors.lastname: George
authors.forename: John
authors.initials: J
authors.lastname: Chen
authors.forename: Tom W
authors.initials: TW
authors.lastname: Muir
authors.forename: Richard P
authors.initials: RP
authors.lastname: Novick
mesh_terms: D019943:Amino Acid Substitution; D001426:Bacterial Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D000071677:Histidine Kinase; D007700:Kinetics; D008024:Ligands; D010456:Peptides, Cyclic; D011485:Protein Binding / Q000502:physiology; D011494:Protein Kinases / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D017433:Protein Structure, Secondary / Q000502:physiology; D017434:Protein Structure, Tertiary / Q000502:physiology; D053038:Quorum Sensing / Q000502:physiology*; D013211:Staphylococcus aureus / Q000201:enzymology* / Q000235:genetics
chemical_list: C414532:AgrD protein, Staphylococcus; D001426:Bacterial Proteins; D008024:Ligands; D010456:Peptides, Cyclic; D011494:Protein Kinases; D000071677:Histidine Kinase