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Antimicrobial Peptide Recognition Motif of the Substrate Binding Protein SapA from Nontypeable Haemophilus influenzae
Kristen G Rivera1, Kari J Tanaka1, Evan R Buechel1
1Department of Molecular Biosciences, Northwestern University, Evanston, Illinois 60208, United States.
Abstract:
Nontypeable Haemophilus influenzae (NTHi) is an opportunistic pathogen associated with respiratory diseases, including otitis media and exacerbations of chronic obstructive pulmonary disease. NTHi exhibits resistance to killing by host antimicrobial peptides (AMPs) mediated by SapA, the substrate binding protein of the sensitivity to antimicrobial peptides (Sap) transporter. However, the specific mechanisms by which SapA selectively binds various AMPs such as defensins and cathelicidin are unknown. In this study, we report mutational analyses of both defensin AMPs and the SapA binding pocket to define the specificity of AMP recognition. Bactericidal assays revealed that NTHi lacking SapA are more susceptible to human beta defensins and LL-37, while remaining highly resistant to a human alpha defensin. In contrast to homologues, our research underscores the distinct specificity of NTHi SapA, which selectively recognizes and binds to peptides containing the charged-hydrophobic motif PKE and RRY. These findings provide valuable insight into the divergence of SapA among bacterial species and NTHi SapA's ability to selectively interact with specific AMPs to mediate resistance.
Insights
Nontypeable Haemophilus influenzae (NTHi) uses SapA to resist antimicrobial peptides (AMPs). This study reveals SapA
Area of Science:
- Microbiology
- Immunology
- Biochemistry
Background:
- Nontypeable Haemophilus influenzae (NTHi) is a key pathogen in respiratory infections.
- NTHi evades host defense via the SapA protein, which confers resistance to antimicrobial peptides (AMPs).
- The precise mechanisms of SapA's selective AMP binding remain unclear.
Purpose of the Study:
- To elucidate the specificity of NTHi SapA in recognizing and binding various AMPs.
- To define the structural basis for NTHi SapA's selective interaction with defensins and cathelicidins.
Main Methods:
- Site-directed mutagenesis of human defensins and the NTHi SapA binding pocket.
- Bactericidal assays to assess NTHi susceptibility to AMPs.
- Analysis of peptide binding specificity based on sequence motifs.
Main Results:
- NTHi strains lacking SapA showed increased susceptibility to human beta defensins and LL-37.
- NTHi remained resistant to a human alpha defensin, indicating selective resistance.
- NTHi SapA specifically recognizes and binds peptides containing the PKE and RRY motifs.
Conclusions:
- NTHi SapA exhibits distinct specificity, differing from homologues in other bacterial species.
- The PKE and RRY motifs are critical for NTHi SapA's selective binding of AMPs.
- Understanding NTHi SapA-AMP interactions provides insights into bacterial resistance mechanisms.
