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. 2023 May 10;13(5):812.
doi: 10.3390/biom13050812.

Structural Basis of PE_PGRS Polymorphism, a Tool for Functional Modulation

Eliza Kramarska  1 , Flavio De Maio  2 , Giovanni Delogu  3   4 , Rita Berisio  1
Affiliations

Structural Basis of PE_PGRS Polymorphism, a Tool for Functional Modulation

Eliza Kramarska et al. Biomolecules. .

Abstract

Background: The mycobacterial PE_PGRS protein family is present only in pathogenic strains of the genus mycobacterium, such as Mtb and members of the MTB complex, suggesting a likely important role of this family in pathogenesis. Their PGRS domains are highly polymorphic and have been suggested to cause antigenic variations and facilitate pathogen survival. The availability of AlphaFold2.0 offered us a unique opportunity to better understand structural and functional properties of these domains and a role of polymorphism in Mtb evolution and dissemination.

Methods: We made extensive use of AlphaFold2.0 computations and coupled them with sequence distribution phylogenetic and frequency analyses, and antigenic predictions.

Results: Modeling of several polymorphic forms of PE_PGRS33, the prototype of the PE_PGRS family and sequence analyses allowed us to predict the structural impact of mutations/deletions/insertions present in the most frequent variants. These analyses well correlate with the observed frequency and with the phenotypic features of the described variants.

Conclusions: Here, we provide a thorough description of structural impacts of the observed polymorphism of PE_PGRS33 protein and we correlate predicted structures to the known fitness of strains containing specific variants. Finally, we also identify protein variants associated with bacterial evolution, showing sophisticated modifications likely endowed with a gain-of-function role during bacterial evolution.

Keywords: PE_PGRS; polymorphism; protein structure; tuberculosis.

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Conflict of interest statement

The authors declare no conflict of interest.

Figures

Figure 1
Figure 1
A schematic view of PE_PGRS33 structure. (A) Cartoon representation of PE_PGRS33 Alphafold2 model. (B) A scheme of domain boundaries and localisation of head and foot residues. The PE domain is represented in purple, the GRPLI motif in blue, head residues in green, foot residues in orange.
Figure 2
Figure 2
Structural features of PGRS33 domain and its GRPLI motif. (A) Surface and cartoon representations of PGRS33 domain (variant of the Mtb H37Rv reference strain). Hydrophobic residues are drawn in stick representation; those belonging to the hydrophobic foot in orange. A representative GGA triplet (308–310) is drawn in blue; the inset shows a detail of hydrogen bonding interactions formed by GGA triplets. (B) Stick representation of the GRPLI motif of PE_PGRS33 and of its interacting residues.
Figure 3
Figure 3
Frequency of the PE_PGRS33 variants as calculated in a collection of 1024 Mtb clinical isolates. PE_PGRS33 variants were functionally classified based on the impact of polymorphisms and indels on aminoacidic sequence. A red dotted line is drawn at 1% frequency.
Figure 4
Figure 4
SNPs on PE_PGRS33 protein. (A) Frequencies of SNPs in PE_PGRS33 and (B) localisation of Gly mutations on the structure of PGRS33allRv. Gly mutations are shown as red (frequency 0.10) and orange (frequency > 0.10) balls. (Frequency%1 in panel A reports pe_pgrs33 allele frequencies as indicated in Supplementary Table S1 and shown in Figure 3).
Figure 5
Figure 5
(A) Cartoon representation of PGRS33all26 (orange) and (B) PGRS33all29 (cyan). The region G139-L161 of PGRS33all26 (panel (A)) deleted in PGRS33all29, is drawn in dark grey. In both variants, a flexible region (white, pLDDT score < 50) originates from the D14 frameshift change. PE_PGRS33all29 also contains the nsSNP P116L, as reported below.
Figure 6
Figure 6
Cartoon representations of the effect of structure-preserving immunomodulating deletions on PGRS33 sandwich fold. Deleted regions are mapped on the wild type PGRS33 in the central panel.
Figure 7
Figure 7
Cartoon representations showing the structural modification predicted by AlphaFold2.0 upon deletion of G196-D243. In the left panel, the three portions of PGRS33allRv 114–195, 196–243 and 244–498 are drawn in grey, black and slate blue, respectively. The same colour code is kept in the right panel, displaying the structure of PGRS33allΔG196-D243. Hydrophobic residues belonging to the foot are drawn in stick representation.
Figure 8
Figure 8
(A) Prediction of B-cell epitopes using BepiPred-3.0 and (B) mapping of most relevant epitopes (score > 0.15) on PGRS33 structure.
Figure 9
Figure 9
Polymorphisms occurring in pe_pgrs33 gene in Mtb complex lineages. (A) Presence of the most relevant pe_pgrs33 polymorphisms detected in Mtb and their association with the main Mtb complex phylogeographic lineages. (B) Graphical representation of the genomic changes in the pe_pgrs33 gene: S, non-synonymous mutations; D, deletions; I, insertions, according to the nomenclature as in [20].
Figure 10
Figure 10
Structure evolution from the most frequent variant of lineage L1 PE_PGRS33all29 (top) to PE_PGRS33AllRv (bottom). The inset in the top panel reports the superposition of GRLPI motifs of the two predicted structures; similar backbone torsion angles are observed for Pro116 and Leu116 (φ = −70, ψ = 150). In the bottom panel, inserted regions from PE_PGRS33all29 to PE_PGRS33Rv variants are drawn in dark blue.
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Authors were funded through the project INF-ACT “One Health Basic and Translational Research Actions addressing Unmet Needs on Emerging Infectious Diseases PE00000007”, PNRR Mission 4, funded by EU “NextGenerationEU”- D.D. MUR Prot.n. 0001554 of 11/10/2022. E.K. was funded by BactiVax—Anti-Bacterial Innovative Vaccines, Marie Skłodowska-Curie Actions, GA 860325.

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