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, 10 (4), e0123249

IMPIPS: The Immune Protection-Inducing Protein Structure Concept in the Search for Steric-Electron and Topochemical Principles for Complete Fully-Protective Chemically Synthesised Vaccine Development


IMPIPS: The Immune Protection-Inducing Protein Structure Concept in the Search for Steric-Electron and Topochemical Principles for Complete Fully-Protective Chemically Synthesised Vaccine Development

Manuel Elkin Patarroyo et al. PLoS One.


Determining immune protection-inducing protein structures (IMPIPS) involves defining the stereo-electron and topochemical characteristics which are essential in MHC-p-TCR complex formation. Modified high activity binding peptides (mHABP) were thus synthesised to produce a large panel of IMPIPS measuring 26.5 ±3.5Å between the farthest atoms fitting into Pockets 1 to 9 of HLA-DRβ1* structures. They displayed a polyproline II-like (PPIIL) structure with their backbone O and N atoms orientated to establish H-bonds with specific residues from HLA-DRβ1*-peptide binding regions (PBR). Residues having specific charge and gauche+ orientation regarding p3χ1, p5χ2, and p7χ1 angles determined appropriate rotamer orientation for perfectly fitting into the TCR to induce an appropriate immune response. Immunological assays in Aotus monkeys involving IMPIPS mixtures led to promising results; taken together with the aforementioned physicochemical principles, non-interfering, long-lasting, protection-inducing, multi-epitope, multistage, minimal subunit-based chemically-synthesised peptides can be designed against diseases scourging humankind.

Conflict of interest statement

Competing Interests: The authors have declared that no competing interests exist.


Fig 1
Fig 1. Representative torsion angles (Φ, Ψ, χ1, χ2, χ3 and χ4) in mHABPs involved in mixtures immunised in Aotus monkeys.
Grey shows the PPIIL region, yellow shows the β-turn region and green an α-helix conformation. Purple box, χ1 angles for p3 and p7, and χ2 angles for p5, highlighting their gauche+ rotamer orientation. The colours of residues vertically displayed in each mHABP correspond to the code for Fig 3.
Fig 2
Fig 2. WB analysis showing the reactivity of Aotus immunised with different mixtures of mHABPs.
A. Reactivity with CSP-derived 25608, 32958, and TRAP 24246 mHABP mixtures 140 days after first immunisation (80 days after 3rd or III80) showing reactivity in 2/8 Aotus with CSP-rI. B. Reactivity with CSP-derived 25608, 32958, and TRAP 24254 and 24246 mHABP mixtures evidencing reactivity with CSP-rII and with TRAP rII in 5/6 Aotus monkeys C. Reactivity with CSP-derived rI 25608, rII 32958, TRAP rII 24254, MSP-2 24112, EBA-175 RI 13790 and rII 24292 and AMA-1 10022.
Fig 3
Fig 3. Spz- and Mrz-derived mHABP fragments binding to the HLA-DRβ1* PBR.
Side view of mHABPs binding to the HLA-DRβ1* PBR (as assessed by 1H-NMR) displaying the residues according to the colour code: p1 (fuchsia), p2 (red), p3 (pale blue), p4 (dark blue), p5 (pink), p6 (orange), p7 (grey), p8 (yellow) and p9 (green). The dotted balls in light green represent the nonbonding free electron pairs able to establish H bonds with the HLA-DRβ1*PBR residues. The pink planes mark peptide bonds.
Fig 4
Fig 4. Comparison of initial and final peptide-bond planes for 25608.37. A, C, E, G, I, K and M.
Green cubes indicate initial theoretical positions for 25068.37 regarding PBR residues forming Φ and ψ angles on planes 1 to 9. B., D., F., H., J., L. and N. Final position of these angles reaching the lowest energy to avoid topochemical steric clashes, as measured for the lowest energy conformer, based on 3D structure obtained by 1H-NMR. Most angles were close to -93° ± 25° (Φ) and 134° ± 15° (Ψ), similar to PPIIL. Minimisation values for atom clashes as determined by Ramachandran plot. HLA-DRβ1* classification according to experimental binding to purified molecules, binding motifs and binding registers is shown at the top on the right-hand side.
Fig 5
Fig 5. Steric-electron effects. 25608.37 (left panel) and 10022.43 (right panel) mHABP residues, displaying hybrid sigma (σ) orbitals (yellow), π and p orbitals perpendicular to them (red, blurred balloons).
For 25608.37: A and B. Phe1 displaying π resonance (red—bonds between p orbitals); Ser2 tetrahedron with the two free electron pairs (indicated) showing only the σ orbitals. C. Leu3 (green), showing the tetrahedron framing Cδ1 side-chain and orientation (pointing upwards), only Gly4 plane is shown. D. Glu5 (green), showing the tetrahedron framing Cɣ and trigon with Cδ and resonance between the two O and their corresponding Cδ from the side-chain (blurred red balloons); the electron charge is shown in blurred red orbitals. E. Asn 6 directed to Pocket 6, showing the tetrahedron, a trigon and the electron charge in blurred red. F. Pro 7 in grey and two trigons in green. G. Asn 8 directed toward the TCR with its corresponding p orbitals and its non-bonding free electron pair; Ala9 is also shown in green with a tetrahedron in the same colour. For 10022.43: H. Phe 1 orientation, π resonance and its planes corresponding to peptide bonds. I. His2 showing the π resonance tiara. J. Pro3 cyclic structure with σ orbitals pointing upwards to contact the TCR, and also displaying the tetrahedron formed by the Ser4 side-chain pointing downwards. K. showing the tetrahedrons formed for Gly5 and Ser7 with its two free electron pairs. L. Pro 8 σ electrons pointing upwards and Val9, showing the two tetrahedrons framing Cδ1 and Cδ2 and their apolarity represented in σ-bonds.
Fig 6
Fig 6. HLA-DRβ1* residues involved in Pocket 1, 4, 6, 9 formation and differences with Aona DRβ.
Molecular surface of amino acids involved in Pocket 1, 4, 6 and 9 formation (α-chain in pink and β-chain in blue). Differences between HLADRβ1* and Aona DR are shown in red. The amino acids forming the pockets are shown on top of each one. A.B.C.D. Spz 25608.37 mHABP (yellow) fitting into HLADRβ1*0404/0401. The βV86 (red) is the dimorphic variant present in Pocket 1 in some HLA-DRβ1* molecules, showing that there were no differences between HLADRβ1*0422 and Aona DRβ3*0603 in this pocket. E. The same held true for HLADRβ1*0301 and Aona DRβ1*0305 in this pocket. G. Aona DRβ1*0305 Fβ9E (in red) replacement in Pocket 6 was located far away from Pocket 6 side wall; it therefore had no impact on mHABP binding when compared to HLADRβ1*0301. H. The 2 replacements observed between Aona DRβ1*0305, HLADRβ1*0301 (Fβ9E and Yβ37N, both in red) had no impact in Pocket 9 since they were located far away from this pocket’s floor. These mHABPs were thus very similar and could be used for human immunisations without any further modifications.
Fig 7
Fig 7. H-bonds established between HLA-DRβ1*0404/0401 with 25608.37 and HLA-DRβ1*0301 with 10022. A.D. Front view B.E. Top view C. F. Side view.
Left panel: 25608.37—HLA-DRβ1*0404/0401. Right-hand Panel: 10022.43—HLA-DRβ1*0301. Their atomic distances are given in S1 Fig.
Fig 8
Fig 8. Ring structure formation in mHABP 25608.37.
H-bonds or vdW interactions between Ser2 25608.37 N and O backbone atoms and HLA-DRβ1*0404/0401 conserved Nβ82 side-chain atoms; this led to establishing ring structures involving the 11 atoms shown here. These 9–11 ring atoms structures were also established with Qα9, Nα62 and Nα69, thereby stabilising IMPIPS binding to the HLA-DRβ1* PBR.
Fig 9
Fig 9. mHABP electron density. Side-chain atoms for p2, p3, p5, p7 and p8 in 25608.37 and 10022.43.
The figure shows the side-chains for upwardly-oriented residues pointing to TCR-contacting residues. Polar amino acids present in p2, p5 and p8, displaying their non-bonding free electron pairs and π orbital surfaces shown in blurred red while σ orbitals for apolar ones present in p3 and p7 are shown by yellow surfaces. The φ angles (≤ -64°) in p7P (25608.37), p5P (32958.2), p7P (24254.31), p3P and p8P (10022.43) oriented this residue to make contact with the TCR.

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Grant support

This work was financed by the “The Colombian Science, Technology and Innovation Department (Colciencias)” (Grant 709-2013). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.