Sub-nanoscale surface ruggedness provides a water-tight seal for exposed regions in soluble protein structure

Abstract

Soluble proteins must maintain backbone hydrogen bonds (BHBs) water-tight to ensure structural integrity. This protection is often achieved by burying the BHBs or wrapping them through intermolecular associations. On the other hand, water has low coordination resilience, with loss of hydrogen-bonding partnerships carrying significant thermodynamic cost. Thus, a core problem in structural biology is whether natural design actually exploits the water coordination stiffness to seal the backbone in regions that are exposed to the solvent. This work explores the molecular design features that make this type of seal operative, focusing on the side-chain arrangements that shield the protein backbone. We show that an efficient sealing is achieved by adapting the sub-nanoscale surface topography to the stringency of water coordination: an exposed BHB may be kept dry if the local concave curvature is small enough to impede formation of the coordination shell of a penetrating water molecule. Examination of an exhaustive database of uncomplexed proteins reveals that exposed BHBs invariably occur within such sub-nanoscale cavities in native folds, while this level of local ruggedness is absent in other regions. By contrast, BHB exposure in misfolded proteins occurs with larger local curvature promoting backbone hydration and consequently, structure disruption. These findings unravel physical constraints fitting a spatially dependent least-action for water coordination, introduce a molecular design concept, and herald the advent of water-tight peptide-based materials with sufficient backbone exposure to remain flexible.

Figure 1. Distances from interfacial water to exposed and buried backbone hydrogen bonds in soluble proteins.

Probability distribution of distances from backbone hydrogen bonds (BHBs, grey line) and exposed backbone hydrogen bonds (EBHBs, black line) to the closest surrounding water molecule. The distributions were obtained from the equilibrium values of the parameter d_min for 377,116 BHBs and 97,120 EBHBs for an exhaustive dataset of 2661 free monomeric proteins reported in PDB (Table S1). The inset displays in virtual bond and space filling representation the EBHB involving residues Asp52-Glu29 of human ubiquitin (PDB.1UBI) with osculating sphere radius θ = 1.7Å.

Figure 2. Surface ruggedness and backbone exposure in soluble proteins.

Probability distribution of sub-nanoscale curvature radii of regions on the protein surface in the vicinity of BHBs (grey line), with the EBHB contribution represented by the black line. A region on the protein surface is defined as being in the vicinity of a BHB if it is shaped by solvent-exposed groups contained within a sphere of radius 3Å centered at the baricenter of the BHB. The curvature radius of a point on the protein water-exposed enveloping surface , is defined as the radius of a first-order contact (osculating) sphere.

Figure 3. Surface curvature dependence of minimum distance and coordination index for water molecule closest to BHB.

Distance to BHB of closest water molecule (d_min in angstroms, grey) and HB-coordination of the molecule (dimensionless g_min, black) plotted as a function of the curvature radius (θ) of the protein surface vicinal to the BHB. The parameters d_min, g_min are computed as averages over all BHBs with the same θ-value. The latter are coarse grained to ¼ of an angstrom. The inset describes the extent of water penetration for three illustrative θ-values.

Figure 4. Distances of exposed BHBs to interfacial water for native folds and misfolded proteins.

Probability distribution of distances from EBHBs of native folds (black line) and from EBHBs of misfolded proteins (grey line) to the closest surrounding water molecule. The datapoints on the grey line were obtained for 25 equilibrated misfolded structures , . The distributions were obtained from the equilibrium values of the parameter d_min for 31,072 BHBs and 17,108 EBHBs from a dataset of misfolded structures obtained by threading a PDB-reported protein onto the structure of another .

Figure 5. Surface ruggedness and backbone exposure for misfolded proteins.

Probability distribution of sub-nanoscale curvature radii of regions on the surface of misfolded proteins in the vicinity of BHBs (grey line), with the EBHB contribution represented by the black line.