Skip to main content
SpringerLink
Log in

Computational study of pH-dependent oligomerization and ligand binding in Alt a 1, a highly allergenic protein with a unique fold

  • Published:
Journal of Computer-Aided Molecular Design Aims and scope Submit manuscript

Abstract

Alt a 1 is a highly allergenic protein from Alternaria fungi responsible for several respiratory diseases. Its crystal structure revealed a unique β-barrel fold that defines a new family exclusive to fungi and forms a symmetrical dimer in a butterfly-like shape as well as tetramers. Its biological function is as yet unknown but its localization in cell wall of Alternaria spores and its interactions in the onset of allergy reactions point to a function to transport ligands. However, at odds with binding features in β-barrel proteins, monomeric Alt a 1 seems unable to harbor ligands because the barrel is too narrow. Tetrameric Alt a 1 is able to bind the flavonoid quercetin, yet the stability of the aggregate and the own ligand binding are pH-dependent. At pH 6.5, which Alt a 1 would meet when secreted by spores in bronchial epithelium, tetramer-quercetin complex is stable. At pH 5.5, which Alt a 1 would meet in apoplast when infecting plants, the complex breaks down. By means of a combined computational study that includes docking calculations, empirical pKa estimates, Poisson–Boltzmann electrostatic potentials, and Molecular Dynamics simulations, we identified a putative binding site at the dimeric interface between subunits in tetramer. We propose an explanation on the pH-dependence of both oligomerization states and protein–ligand affinity of Alt a 1 in terms of electrostatic variations associated to distinct protonation states at different pHs. The uniqueness of this singular protein can thus be tracked in the combination of all these features.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Fig. 1
Fig. 2
Fig. 3
Fig. 4
Fig. 5
Fig. 6
Fig. 7
Fig. 8

Similar content being viewed by others

Abbreviations

3D:

Three-dimensional

APBS:

Adaptive Poisson–Boltzmann solver

DPBA:

Diphenylboric acid 2-aminoethyl ester

MD:

Molecular dynamics

PB:

Poisson–Boltzmann

PDB:

Protein data bank

PISA:

Proteins, interfaces, structures, and assemblies

PTGL:

Protein topology graph library

RMSD:

Root mean square deviation

RMSF:

Root mean square fluctuation

References

  1. Delfino RJ, Zeiger RS, Seltzer JM, Street DH, Matteucci RM, Anderson PR et al (1997) The effect of outdoor fungal spore concentrations on daily asthma severity. Environ Health Perspect 105:622–635

    Article  CAS  Google Scholar 

  2. Bush RK, Portnoy JM (2001) The role and abatement of fungal allergens in allergic diseases. J Allergy Clin Immunol 107:S430–S440

    Article  CAS  Google Scholar 

  3. Anderson M, Downs S, Mitakatis T, Leuppi J, Marks G (2003) Natural exposure to Alternaria spores induces allergic rhinitis symptoms in sensitized children. Pediatr Allergy Immunol 14:100–105

    Article  Google Scholar 

  4. Bush RK, Prochnau JJ (2004) Alternaria-induced asthma. J Allergy Clin Immunol 113:227–234

    Article  Google Scholar 

  5. Knutsen AP, Bush RK, Demain JG, Denning DW, Dixit A, Fairs A et al (2012) Fungi and allergic lower respiratory tract diseases. J Allergy Clin Immunol 129:280–289

    Article  Google Scholar 

  6. Salo PM, Arbes SJ, Sever M, Jaramillo R, Cohn RD, London SJ, Zeldin DC (2006) Exposure to Alternaria alternata in US homes is associated with asthma symptoms. J Allergy Clin Immunol 118:892–898

    Article  Google Scholar 

  7. Feo Brito F, Alonso AM, Carnes J, Martin-Martin R, Fernandez-Caldas E, Galindo PA et al (2012) Correlation between Alt a 1 levels and clinical symptoms in Alternaria alternata-monosensitized patients. J Invest Allergol Clin Immunol 22:154–159

    CAS  Google Scholar 

  8. Deards MJ, Montague AE (1991) Purification and characterization of a major allergen of Alternaria alternata. Mol Immunol 28:409–415

    Article  CAS  Google Scholar 

  9. De Vouge MW, Thaker AJ, Curran IH, Zhang L, Muradia G, Rode H et al (1996) Isolation and expression of a cDNA clone encoding an Alternaria alternata Alt a 1 subunit. Int Arch Allergy Immunol 111:385–395

    Article  Google Scholar 

  10. Vailes LD, Perzanowski MS, Wheatley LM, Platt-Mills TA, Chapman MD (2001) IgE and IgG antibody responses to recombinant Alt a 1 as a marker of sensitization to Alternaria in asthma and atopic dermatitis. Clin Exp Allergy 31:1891–1895

    Article  CAS  Google Scholar 

  11. Asturias JA, Ibarrola I, Ferrer A, Andreu C, Lopez-Pascual E, Quiralte J et al (2005) Diagnosis of Alternaria alternata sensitization with natural and recombinant Alt a 1 allergens. J Allergy Clin Immunol 115:1210–1217

    Article  CAS  Google Scholar 

  12. Twaroch TE, Focke M, Fleischmann K, Balic N, Lupinek K, Blatt K et al (2012) Carrier-bound Alt a 1 peptides without allergenic activity for vaccination against Alternaria alternata allergy. Clin Exp Allergy 42:966–975

    CAS  Google Scholar 

  13. Kurup VP, Vijay HM, Kumar V, Castillo L, Elms N (2003) IgE binding synthetic peptides of Alt a 1, a major allergen of Alternaria alternata. Peptides 24:179–185

    Article  CAS  Google Scholar 

  14. Chrusczc M, Chapman MD, Osinski T, Solberg R, Demas M, Porebski PJ et al (2012) Alternaria alternata allergen Alt a 1: a unique β-barrel protein dimer found exclusively in fungi. J Allergy Clin Immunol 130:241–247

    Article  Google Scholar 

  15. Rouvinen J, Janis J, Laukhanen ML, Jylha S, Niemi M, Paivinen T et al (2010) Transient dimers of allergens. PLoS One 5:e9037/1-9

    Article  Google Scholar 

  16. Wagner GE, Gutfreund S, Fauland K, Keller W, Valenta R, Zangger K (2014) Backbone resonance assignment of Alt a 1, a unique β-barrel protein and the major allergen of Alternaria alternata. Biomol NMR Assign 8:229–231

    Article  CAS  Google Scholar 

  17. Twaroch TE, Arcalis E, Sterflinger K, Stoger E, Swoboda I, Valenta R (2012) Predominant localization of the major Alternaria allergen Alt a 1 in the cell wall of airborne spores. J Allergy Clin Immunol 129:1148–1149

    Article  CAS  Google Scholar 

  18. Mitakakis TZ, Barnes C, Tovey ER (2011) Spore germination increases allergen release from Alternaria. J Allergy Clin Immunol 107:388–390

    Article  Google Scholar 

  19. Garrido-Arandia M, Gómez-Casado C, Díaz-Perales A, Pacios LF (2014) Molecular dynamics of major allergens from Alternaria, birch pollen and peach. Mol Inf 33:682–694

    Article  CAS  Google Scholar 

  20. Kabsch W, Sander C (1983) Dictionary of protein secondary structure: pattern recognition of hydrogen-bonded and geometrical features. Biopolymers 22:2577–2637

    Article  CAS  Google Scholar 

  21. Touw WG, Baakman C, Black J, Te Beek TAH, Krieger E, Robbie P et al (2015) A series of PDB related databases for everyday needs. Nucleic Acids Res 43:D364–D368

    Article  Google Scholar 

  22. Laskowski RA (2001) PDBsum: summaries and analyses of PDB structures. Nucleic Acids Res 29:221–222

    Article  CAS  Google Scholar 

  23. Laskowski RA (2014) PDBsum additions. Nucleic Acids Res 37:D355–D359

    Article  Google Scholar 

  24. May P, Barthel S, Koch I (2004) Protein topology graph library. Bioinformatics 20:3277–3279

    Article  CAS  Google Scholar 

  25. Krissinel E, Henrick K (2007) Inference of macromolecular assemblies from crystalline state. J Mol Biol 372:774–797

    Article  CAS  Google Scholar 

  26. Krissinel E (2010) Crystal contacts as nature’s docking solutions. J Comput Chem 31:133–143

    Article  CAS  Google Scholar 

  27. The PyMOL Molecular Graphics System, Version 1.7.6.4 Schrödinger, LLC

  28. Pettersen EF, Goddard TD, Huang CC, Couch GS, Greenblatt DM, Meng EC, Ferrin TE (2004) UCSF Chimera: a visualization system for exploratory research and analysis. J Comput Chem 25:1605–1612

    Article  CAS  Google Scholar 

  29. Siegel D, Troyanov S, Noack J, Emmerling F, Nehls I (2010) Alternariol. Acta Crystallogr Sect E 66:1366/1-7

    Article  Google Scholar 

  30. Maier JA, Martinez C, Kasavajhala K, Wickstrom L, Hauser KE, Simmerling C (2015) ff14SB: improving the accuracy of protein side chain and backbone parameters from ff99SB. J Chem Theor Comput 11:3696–3713

    Article  CAS  Google Scholar 

  31. Wang J, Wang W, Kollman PA, Case DA (2006) Automatic atom type and bond type perception in molecular mechanical calculations. J Mol Graph Model 25:247–260

    Article  Google Scholar 

  32. Morris GM, Huey R, Lindstrom W, Sanner MF, Belew RK, Goodsell DS, Olson AJ (2009) Autodock4 and AutoDockTools4: automated docking with selective receptor flexibility. J Comput Chem 16:2785–2791

    Article  Google Scholar 

  33. Trott O, Olson AJ (2009) AutoDock Vina: Improving the speed and accuracy of docking with a new scoring function, efficient optimization, and multithreading. J Comput Chem 31:455–461

    Google Scholar 

  34. Olsson MHM, Sondergaard CR, Rostkowski M, Jensen JH (2011) PROPKA3: Consistent treatment of internal and surface residues in empirical pKa predictions. J Chem Theor Comput 7:525–537

    Article  CAS  Google Scholar 

  35. Sondergaard CR, Olsson MHM, Rostkowski M, Jensen JH (2011) Improved treatment of ligands and coupling effects in empirical calculation and rationalization of pKa values. J Chem Theor Comput 7:2284–2295

    Article  CAS  Google Scholar 

  36. Dolinsky TJ, Nielsen JE, McCammon JA, Baker NA (2004) PDB2PQR: an automated pipeline for the setup, execution, and analysis of Poisson-Boltzmann electrostatics calculations. Nucl Acids Res 32:W665–W667

    Article  CAS  Google Scholar 

  37. Dolinsky TJ, Czodrowski P, Li H, Nielsen JE, Jensen JH, Klebe G, Baker NA (2007) PDB2PQR: expanding and upgrading automated preparation of biomolecular structures for molecular simulations. Nucl Acids Res 35:W522–W525

    Article  Google Scholar 

  38. MacKerell A, Bashford D, Bellott M, Dunbrack RL, Evanseck J, Field MJ et al (1998) All-atom empirical potential for molecular modeling and dynamics studies of proteins. J Phys Chem B 102:3586–3616

    Article  CAS  Google Scholar 

  39. Baker NA, Sept D, Joseph S, Holst MJ, McCammon JA (2001) Electrostatics of nanosystems: application to microtubules and the ribosome. Proc Natl Acad Sci USA 98:10037–10041

    Article  CAS  Google Scholar 

  40. Zoete V, Cuendet MA, Grosdidier A, Michielin O (2011) SwissParam, a fast force field generation tool for small organic molecules. J Comput Chem 32:2359–2368

    Article  CAS  Google Scholar 

  41. Phillips JC, Braun R, Wang W, Gumbart J, Tajkhorshid E, Villa E et al (2005) Scalable molecular dynamics with NAMD. J Comput Chem 26:1781–1802

    Article  CAS  Google Scholar 

  42. Jorgensen WL, Chandrasekhar J, Madura JD, Impey RW, Klein ML (1983) Comparison of simple potential functions for simulating liquid water. J Chem Phys 79:926–935

    Article  CAS  Google Scholar 

  43. Darden T, York D, Pedersen L (1993) Particle mesh Ewald: an N log(N) method for Ewald sums in large systems. J Chem Phys 98:10089–10092

    Article  CAS  Google Scholar 

  44. Humphrey W, Dalke A, Schulten K (1996) VMD—visual molecular dynamics. J Mol Graph 14:33–38

    Article  CAS  Google Scholar 

  45. Fischer F, Wissicombe JH (2006) Mechanism of acid and base secretion by the airway epithelium. J Membr Biol 211:139–150

    Article  CAS  Google Scholar 

  46. Almeida DPF, Huber DJ (1999) Apoplastic pH and inorganic ion levels in tomato fruit: a potential means for regulation of cell wall metabolism during ripening. Physiol Plant 105:506–512

    Article  CAS  Google Scholar 

  47. Cianci M, Folli C, Zonta F, Florio P, Berni R, Zanotti G (2015) Structural evidence for asymmetric ligand binding to transthyretin. Acta Crystallogr Sect D 71:1582–1592

    Article  CAS  Google Scholar 

  48. Pasquato N, Berni R, Folli C, Alfieri B, Cendron L, Zanotti G (2007) Acidic pH-induced conformational change in amyloidogenic mutant transthyretin. J Mol Biol 366:711–719

    Article  CAS  Google Scholar 

  49. Barkai-Golan R (2008) Alternaria mycotoxins. In: Barkai-Golan R, Nachman P (eds) Mycotoxins in fruits and vegetables. Academic Press, San Diego, pp 185–203

    Chapter  Google Scholar 

  50. European Food Safety Authority (2011) Scientific opinion on the risks for animal and public health related to the presence of Alternaria toxins in feed and food. EFSA J 9(2407):1–97

    Google Scholar 

  51. Garrido-Arandia M, Gómez-Casado C, Díaz-Perales A, Pacios LF (2014) Distortion from planarity in arenes produced by internal rotation of one single hydroxyl hydrogen: the case of alternariol. J Mol Graph Model 53:140–147

    Article  CAS  Google Scholar 

Download references

Acknowledgments

All Molecular Dynamics calculations were carried out on the Magerit supercomputer of Universidad Politécnica de Madrid. The authors acknowledge the computer resources and technical assistance provided by the Centro de Supercomputación y Visualización de Madrid (CeSViMa). This work was supported by the Spanish Ministerio de Ciencia e Innovación MINECO (Grant BIO2013-41403-R) and Instituto de Salud Carlos III, Spain, RETICS 2007 (RD12/0013/14). CGC was supported by the FPI programme from Spanish Government (MICINN/MINECO, Grant BES-2010-034628).

Author information

Authors and Affiliations

  1. Department of Biotechnology and Centre for Plant Genomics and Biotechnology (CBGP), Technical University of Madrid, Pozuelo de Alarcon, 28223, Madrid, Spain

    María Garrido-Arandia, Jorge Bretones, Cristina Gómez-Casado, Nuria Cubells & Araceli Díaz-Perales

  2. Immunology Section, Department of Experimental Medical Sciences, Lund University, BMC D14, 22184, Lund, Sweden

    Cristina Gómez-Casado

  3. Department of Natural Systems and Resources and Centre for Plant Genomics and Biotechnology (CBGP), Technical University of Madrid, 28040, Madrid, Spain

    Luis F. Pacios

Authors
  1. María Garrido-Arandia
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Jorge Bretones
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Cristina Gómez-Casado
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Nuria Cubells
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Araceli Díaz-Perales
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. Luis F. Pacios
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Luis F. Pacios.

Electronic supplementary material

Below is the link to the electronic supplementary material.

Supplementary material 1 (DOCX 2621 kb)

Rights and permissions

Reprints and permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

Garrido-Arandia, M., Bretones, J., Gómez-Casado, C. et al. Computational study of pH-dependent oligomerization and ligand binding in Alt a 1, a highly allergenic protein with a unique fold. J Comput Aided Mol Des 30, 365–379 (2016). https://doi.org/10.1007/s10822-016-9911-6

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s10822-016-9911-6

Keywords

Search

Navigation