Molecular Structural Assignment of Wide-Angle X-ray Scattering for Type I Tropocollagen in Aqueous Solution

Ying-Jen Shiu^1*, Bradley W. Mansel^1,2, Kuei-Fen Liao¹, Ting-Wei Hsu¹, Je-Wei Chang¹, Orion Shih¹, Yi-Qi Yeh¹, U-Ser Jeng^1,3

¹Soft Matter Science Group, TPS13A, National Synchrotron Radiation Research Center, Hsinchu, Taiwan
²Fonterra Research and Development Centre, Fonterra, Palmerston North, New Zealand
³Department of Chemical Engineering & College of Semiconductor Research, National Tsing Hua University, Hsinchu, Taiwan

* Presenter:Ying-Jen Shiu, email:yingjenshiu@gmail.com

The distribution of hydration water, characterized by its heterogeneity, plays a crucial role in governing the arrangement of biomolecules, such as the assembly of collagen into the structural framework of tissues like bones, tendons, and dermis.[1] Despite the utilization of wide-angle X-ray scattering (WAXS), our current understanding of the hydration structure with specific protein surfaces remains insufficient. In this investigation, we aim to illustrate the molecular structure of the hydration water at interfaces and the Type-I tropocollagen using both WAXS measurement [2] and molecular dynamics (MD) simulations. We optimized the structure of truncated collagen, featuring a contour length of 13 nm and 51 amino acids per single chain, enveloped by a 7 Å-thick water layer, through MD simulations and calculated the SWAX within the q range of 0.06 -2.5 Å^-1 using the GROMACS-SWAXS software.[3] Our MD-SWAXS calculation yielded results consistent with experimental WAXS profiles of tropocollagen in a pH = 2 solution, as shown in Figure 1. We also compared the WAXS of collagen without water using the Debye equation. Our MD-SWAXS model reveals that the hydration layer, approximately 2.7 Å - 2.9 Å thick, contributes significantly to the scattering band q0 centered at 1.57 Å^-1. This contribution primarily arises from the molecular structure of collagen and the orientation of water molecules, which form hydrogen bonds with the polar carbonyl groups of collagens. The first layer of the hydrogen bonding network extends to 4.0 Å, encompassing the interfacial surface of collagen, and is anchored by the interaction between hydration water molecules and carboxyl groups. In addition, we employed radial distribution function (RDF) analysis, based on the MD-optimized collagen structure in the aqueous solution, to identify the characteristic distances corresponding to the WAXS patterns. We found that the q1 spike corresponds to atomic spans of 2.8 Å, 3.16 Å, and 2.84 Å, representing the d-spacings between two neighbored peptide units (N-Cα-C-O) within the collagen intra-chains, as well as between collagen and hydration water molecules. Scattering bands q2 and q3 are associated with ordered atomic distances within inter-chain collagen structures, including the d-spacing of C--C, N--N, O--O, Cα--Cα.

Keywords: Tropocollagen, SWAXS, worm-like model, molecular dynamics simulation, hydration shell