Animal Protection and Structural Studies of a Consensus Sequence Vaccine Targeting the Receptor Binding Domain of the Type IV Pilus of Pseudomonas aeruginosa

Abstract Background Tracking the genetic variability of Severe Acute Respiratory Syndrome CoronaVirus 2 (SARS-CoV-2) is a crucial challenge. Mainly to identify target sequences in order to generate robust vaccines and neutralizing monoclonal antibodies, but also to track viral genetic temporal and geographic evolution and to mine for variants associated with reduced or increased disease severity. Several online tools and bioinformatic phylogenetic analyses have been released, but the main interest lies in the Spike protein, which is the pivotal element of current vaccine design, and in the Receptor Binding Domain, that accounts for most of the neutralizing the antibody activity. Methods Here, we present an open-source bioinformatic protocol, and a web portal focused on SARS-CoV-2 single mutations and minimal consensus sequence building as a companion vaccine design tool. Furthermore, we provide immunogenomic analyses to understand the impact of the most frequent RBD variations. Results Results on the whole GISAID sequence dataset at the time of the writing (October 2020) reveals an emerging mutation, S477N, located on the central part of the Spike protein Receptor Binding Domain, the Receptor Binding Motif. Immunogenomic analyses revealed some variation in mutated epitope MHC compatibility, T-cell recognition, and B-cell epitope probability for most frequent human HLAs. Conclusions This work provides a framework able to track down SARS-CoV-2 genomic variability.

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NMR solution structure of the receptor binding domain of Pseudomonas aeruginosa pilin strain P1. Identification of a β-turn

International journal of peptide & protein research ◽

10.1111/j.1399-3011.1996.tb00873.x ◽

2009 ◽

Vol 48 (6) ◽

pp. 539-552 ◽

Cited By ~ 10

Author(s):

A. PATRICIA CAMPBELL ◽

HASMUKH SHETH ◽

ROBERT S. HODGES ◽

BRIAN D. SYKES

Keyword(s):

Pseudomonas Aeruginosa ◽

Receptor Binding ◽

Solution Structure ◽

Binding Domain ◽

Receptor Binding Domain ◽

Nmr Solution Structure

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Pore-forming pyocin S5 utilizes the FptA ferripyochelin receptor to kill Pseudomonas aeruginosa

Microbiology ◽

10.1099/mic.0.070672-0 ◽

2014 ◽

Vol 160 (2) ◽

pp. 261-269 ◽

Cited By ~ 28

Author(s):

Ameer Elfarash ◽

Jozef Dingemans ◽

Lumeng Ye ◽

Ahmed Amir Hassan ◽

Michael Craggs ◽

...

Keyword(s):

Pseudomonas Aeruginosa ◽

Receptor Binding ◽

Protein Product ◽

Binding Domain ◽

Burkholderia Cenocepacia ◽

Receptor Binding Domain ◽

Amino Acid Residues ◽

Gene Encoding ◽

Siderophore Receptors ◽

Transposon Mutants

Pyocins are toxic proteins produced by some strains of Pseudomonas aeruginosa that are lethal for related strains of the same species. Some soluble pyocins (S2, S3 and S4) were previously shown to use the pyoverdine siderophore receptors to enter the cell. The P. aeruginosa PAO1 pore-forming pyocin S5 encoding gene (PAO985) was cloned into the expression vector pET15b, and the affinity-purified protein product tested for its killing activity against different P. aeruginosa strains. The results, however, did not show any correlation with a specific ferripyoverdine receptor. To further identify the S5 receptor, transposon mutants were generated. Pooled mutants were exposed to pyocin S5 and the resistant colonies growing in the killing zone were selected. The majority of S5-resistant mutants had an insertion in the fptA gene encoding the receptor for the siderophore pyochelin. Complementation of an fptA transposon mutant with the P. aeruginosa fptA gene in trans restored the sensitivity to S5. In order to define the receptor-binding domain of pyocin S5, two hybrid pyocins were constructed containing different regions from pyocin S5 fused to the C-terminal translocation and DNase killing domains of pyocin S2. Only the protein containing amino acid residues 151 to 300 from S5 showed toxicity, indicating that the pyocin S5 receptor-binding domain is not at the N-terminus of the protein as in other S-type pyocins. Pyocin S5 was, however, unable to kill Burkholderia cenocepacia strains producing a ferripyochelin FptA receptor, nor was the B. cenocepacia fptA gene able to restore the sensitivity of the resistant fptA mutant P. aeruginosa strain.

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Use of synthetic peptides to confirm that the Pseudomonas aeruginosa PAK pilus adhesin and the Candida albicans fimbrial adhesin possess a homologous receptor‐binding domain

Molecular Microbiology ◽

10.1046/j.1365-2958.1996.454982.x ◽

1996 ◽

Vol 19 (5) ◽

pp. 1107-1116 ◽

Cited By ~ 18

Author(s):

Lei Yu ◽

Kok K. Lee ◽

William Paranchych ◽

Robert S. Hodges ◽

Randall T. Irvin

Keyword(s):

Pseudomonas Aeruginosa ◽

Candida Albicans ◽

Receptor Binding ◽

Synthetic Peptides ◽

Binding Domain ◽

Receptor Binding Domain ◽

Fimbrial Adhesin

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A Colicin M-Type Bacteriocin fromPseudomonas aeruginosaTargeting the HxuC Heme Receptor Requires a Novel Immunity Partner

Applied and Environmental Microbiology ◽

10.1128/aem.00716-18 ◽

2018 ◽

Vol 84 (18) ◽

Cited By ~ 6

Author(s):

Maarten G. K. Ghequire ◽

Başak Öztürk

Keyword(s):

Pseudomonas Aeruginosa ◽

Membrane Protein ◽

Receptor Binding ◽

Binding Domain ◽

Modular Organization ◽

Receptor Binding Domain ◽

Inner Membrane ◽

Content Type ◽

Transmembrane Topology ◽

Inner Membrane Protein

ABSTRACTPyocins are bacteriocins secreted byPseudomonas aeruginosa, and they assist in the colonization of different niches. A major subset of these antibacterial proteins adopt a modular organization characteristic of polymorphic toxins. They include a receptor-binding domain, a segment enabling membrane passage, and a toxin module at the carboxy terminus, which eventually kills the target cells. To protect themselves from their own products, bacteriocin-producing strains express an immunity gene concomitantly with the bacteriocin. We show here that a pyocin equipped with a phylogenetically distinct ColM toxin domain, PaeM4, mediates antagonism against a large set ofP. aeruginosaisolates. Immunity to PaeM4 is provided by the inner membrane protein PmiC, which is equipped with a transmembrane topology not previously described for the ColM family. Given that strains lacking apmiCgene are killed by PaeM4, the presence of such an immunity partner likely is a key criterion for escaping cellular death mediated by PaeM4. The presence of a TonB box in PaeM4 and enhanced bacteriocin activity under iron-poor conditions strongly suggested the targeting of a TonB-dependent receptor. Evaluation of PaeM4 activities against TonB-dependent receptor knockout mutants inP. aeruginosaPAO1 revealed that the heme receptor HxuC (PA1302) serves as a PaeM4 target at the cellular surface. Because other ColM-type pyocins may target the ferrichrome receptor FiuA, our results illustrate the versatility in target recognition conferred by the polymorphic nature of ColM-type bacteriocins.IMPORTANCEThe antimicrobial armamentarium of a bacterium is a major asset for colonizing competitive environments. Bacteriocins comprise a subset of these compounds. Pyocins are an example of such antibacterial proteins produced byPseudomonas aeruginosa, killing otherP. aeruginosastrains. A large group of these molecules show a modular protein architecture that includes a receptor-binding domain for initial target cell attachment and a killer domain. In this study, we have shown that a novel modular pyocin (PaeM4) that kills target bacteria via interference with peptidoglycan assembly takes advantage of the HxuC heme receptor. Cells can protect themselves from killing by the presence of a dedicated immunity partner, an integral inner membrane protein that adopts a transmembrane topology distinct from that of proteins currently known to provide immunity against such toxin activity. Understanding the receptors with which pyocins interact and how immunity to pyocins is achieved is a pivotal step toward the rational design of bacteriocin cocktails for the treatment ofP. aeruginosainfections.

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Adherence of Pseudomonas aeruginosa and Candida albicans to glycosphingolipid (Asialo-GM1) receptors is achieved by a conserved receptor-binding domain present on their adhesins.

Infection and Immunity ◽

10.1128/iai.62.12.5213-5219.1994 ◽

1994 ◽

Vol 62 (12) ◽

pp. 5213-5219 ◽

Cited By ~ 26

Author(s):

L Yu ◽

K K Lee ◽

R S Hodges ◽

W Paranchych ◽

R T Irvin

Keyword(s):

Pseudomonas Aeruginosa ◽

Candida Albicans ◽

Receptor Binding ◽

Binding Domain ◽

Receptor Binding Domain ◽

Asialo Gm1

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NMR solution structure and flexibility of a peptide antigen representing the receptor binding domain of Pseudomonas aeruginosa

Biochemistry ◽

10.1021/bi00212a008 ◽

1993 ◽

Vol 32 (49) ◽

pp. 13432-13440 ◽

Cited By ~ 25

Author(s):

Campbell McInnes ◽

Frank D. Sonnichsen ◽

Cyril M. Kay ◽

Robert S. Hodges ◽

Brian D. Sykes

Keyword(s):

Pseudomonas Aeruginosa ◽

Receptor Binding ◽

Solution Structure ◽

Binding Domain ◽

Receptor Binding Domain ◽

Peptide Antigen ◽

Nmr Solution Structure

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Design of a Companion Bioinformatic Tool to detect the emergence and geographical distribution of SARS-CoV-2 Spike protein genetic variants

10.1101/2020.06.22.133355 ◽

2020 ◽

Author(s):

Alice Massacci ◽

Eleonora Sperandio ◽

Lorenzo D’Ambrosio ◽

Mariano Maffei ◽

Fabio Palombo ◽

...

Keyword(s):

Receptor Binding ◽

Consensus Sequence ◽

Cell Epitope ◽

Vaccine Design ◽

Binding Domain ◽

Spike Protein ◽

Antibody Activity ◽

Binding Motif ◽

Receptor Binding Domain ◽

The Impact

AbstractBackgroundTracking the genetic variability of Severe Acute Respiratory Syndrome CoronaVirus 2 (SARS-CoV-2) is a crucial challenge. Mainly to identify target sequences in order to generate robust vaccines and neutralizing monoclonal antibodies, but also to track viral genetic temporal and geographic evolution and to mine for variants associated with reduced or increased disease severity. Several online tools and bioinformatic phylogenetic analyses have been released, but the main interest lies in the Spike protein, which is the pivotal element of current vaccine design, and in the Receptor Binding Domain, that accounts for most of the neutralizing the antibody activity.MethodsHere, we present an open-source bioinformatic protocol, and a web portal focused on SARS-CoV-2 single mutations and minimal consensus sequence building as a companion vaccine design tool. Furthermore, we provide immunogenomic analyses to understand the impact of the most frequent RBD variations.ResultsResults on the whole GISAID sequence dataset at the time of the writing (October 2020) reveals an emerging mutation, S477N, located on the central part of the Spike protein Receptor Binding Domain, the Receptor Binding Motif. Immunogenomic analyses revealed some variation in mutated epitope MHC compatibility, T-cell recognition, and B-cell epitope probability for most frequent human HLAs.ConclusionsThis work provides a framework able to track down SARS-CoV-2 genomic variability.

Download Full-text