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Journal of Biological Physics

, Volume 38, Issue 4, pp 681–703 | Cite as

Folding of pig gastric mucin non-glycosylated domains: a discrete molecular dynamics study

  • Bogdan Barz
  • Bradley S. Turner
  • Rama Bansil
  • Brigita UrbancEmail author
Original Paper
First Online:

Abstract

Mucin glycoproteins consist of tandem-repeating glycosylated regions flanked by non-repetitive protein domains with little glycosylation. These non-repetitive domains are involved in polymerization of mucin and play an important role in the pH-dependent gelation of gastric mucin, which is essential for protecting the stomach from autodigestion. We examine folding of the non-repetitive sequence of PGM-2X (242 amino acids) and the von Willebrand factor vWF-C1 domain (67 amino acids) at neutral and low pH using discrete molecular dynamics (DMD) in an implicit solvent combined with a four-bead peptide model. Using the same implicit solvent parameters, folding of both domains is simulated at neutral and low pH. In contrast to vWF-C1, PGM-2X folding is strongly affected by pH as indicated by changes in the contact order, radius of gyration, free-energy landscape, and the secondary structure. Whereas the free-energy landscape of vWF-C1 shows a single minimum at both neutral and low pH, the free-energy landscape of PGM-2X is characterized by multiple minima that are more numerous and shallower at low pH. Detailed structural analysis shows that PGM-2X partially unfolds at low pH. This partial unfolding is facilitated by the C-terminal region GLU236-PRO242, which loses contact with the rest of the domain due to effective “mean-field” repulsion among highly positively charged N- and C-terminal regions. Consequently, at low pH, hydrophobic amino acids are more exposed to the solvent. In vWF-C1, low pH induces some structural changes, including an increased exposure of CYS at position 67, but these changes are small compared to those found in PGM-2X. For PGM-2X, the DMD-derived average β-strand propensity increases from 0.26 ± 0.01 at neutral pH to 0.38 ± 0.01 at low pH. For vWF-C1, the DMD-derived average β-strand propensity is 0.32 ± 0.02 at neutral pH and 0.35 ± 0.02 at low pH. The DMD-derived structural information provides insight into pH-induced changes in the folding of two distinct mucin domains and suggests plausible mechanisms of the aggregation/gelation of mucin.

Keywords

Mucin PGM-2X vWF-C1 Mucin gelation DMD simulation Protein folding Free-energy landscape Pig gastric mucin 
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Notes

Acknowledgements

The authors thank Dr. Yuriy V. Sereda for his contribution to the implementation of PRO amino acid into the four-bead protein model. B.U. and B.B. acknowledge the support by the NIH grant AG027818 and thank NSF for the access to the Extreme Science and Engineering Discovery Environment (XSEDE) supercomputing facilities through the grant PHYS100030. B.S.T. thanks Dr. Nezam Afdhal, M.D., Beth Israel Deaconess Medical Center for financial support.

Supplementary material

10867_2012_9280_MOESM1_ESM.eps (4.2 mb)
Figure S1 Normalized distributions of (a) the distance between the Cα atoms of a PRO residue and the residue directly preceding it in the sequence and (b) the dihedral angle ω defining trans (ω=π) versus cis (ω=0) conformation. The distributions were calculated using 15,000 reported protein structures from the Protein Data Bank (EPS 4.24 MB)
10867_2012_9280_MOESM2_ESM.eps (3.3 mb)
Figure S2 Normalized distributions of pair-wise RMSD values for all initial and final DMD-derived conformations of (a) PGM-2X and (b) vWF-C1 domains. Distributions for final conformations at neutral and low pH are shown in black and red, respectively (EPS 3.31 MB)
10867_2012_9280_MOESM3_ESM.tiff (625 kb)
Figure S3 A difference between the neutral and low pH contact map of PGM-2X folded structures (see Fig. 5). The triangle below the diagonal shows positive values of the contact map difference, i.e., the contacts that are stronger at neutral pH and the triangle above the diagonal shows the negative values of the contact map difference, i.e., the contacts that are stronger at low pH. The color scale quantifying the average number of contacts between two residues is displayed on the right (TIFF 624 kb)
10867_2012_9280_MOESM4_ESM.tiff (513 kb)
Figure S4 A difference between the neutral and low pH contact map of vWF-C1 folded structures (see Fig. 10). The triangle below the diagonal shows positive values of the contact map difference, i.e., the contacts that are stronger at neutral pH and the triangle above the diagonal shows the negative values of the contact map difference, i.e., the contacts that are stronger at low pH. The color scale quantifying the average number of contacts between two residues is displayed on the right (TIFF 513 kb)
10867_2012_9280_MOESM5_ESM.eps (3.7 mb)
Figure S5 Probability distribution of the separation |i-j| along the protein sequence between CYS residue i and CYS residue j for (A) PGM-2X and (B) vWF-C1 folded structures. Only the CYS-CYS residue pairs with respective Cβ atoms within a distance of 0.42 nm, corresponding to disulfide bonds in the model, were considered. Distributions at neutral and low pH conformations are shown in black and red, respectively (EPS 3.69 MB)
10867_2012_9280_MOESM6_ESM.tiff (947 kb)
Figure S6 PGM-2X protein models predicted by the I-TASSER server. The N-terminal amino acid ASP1 is colored red and the C-terminal amino acid PRO242 is colored blue. The images were generated by the VMD software package [25] (TIFF 946 kb)
10867_2012_9280_MOESM7_ESM.tiff (672 kb)
Figure S7 vWF-C1 protein models predicted by the I-TASSER server. The N-terminal amino acid CYS1 is colored red and the C-terminal amino acid CYS67 is colored blue. The images were generated by the VMD software package [25] (TIFF 671 kb)

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Copyright information

© Springer Science+Business Media B.V. 2012

Authors and Affiliations

  • Bogdan Barz
    • 1
  • Bradley S. Turner
    • 2
  • Rama Bansil
    • 3
  • Brigita Urbanc
    • 1
    Email author
  1. 1.Physics Dept.Drexel UniversityPhiladelphiaUSA
  2. 2.Division of GastroenterologyBeth Israel Deaconess Medical Center and Harvard Medical SchoolBostonUSA
  3. 3.Physics Dept. and Center for Polymer StudiesBoston UniversityBostonUSA

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