Molecular dynamics reveals how calcium drives hetero- versus homodimerization of type I collagen.

Type I collagen is the main structural protein of vertebrates and forms molecular trimers from the COL1A1 and COL1A2 gene products, proα1(I) and proα2(I), during biosynthesis. Calcium ions are required for trimers to form. The amino acid sequence of the C-propeptide of collagen, which is removed before collagen fibril formation, initially drives heterotrimerization. Abnormal homotrimeric type I collagen is associated with age-related diseases including cancer, fibrosis, and musculoskeletal and cardiovascular conditions, but the circumstances under which the homotrimer may form are poorly understood. Here, we used molecular dynamics simulations of the C-propeptide protein structure to show that inter- and intrachain hydrogen bonding is affected by loss of calcium and that this leads chains to become destabilized, particularly at the interfaces of each chain. Loss of calcium resulted in increased distances between the cysteine residues that form interchain disulfide bonds, preventing the formation of these bonds. Pulling simulations and modeling of calcium dissociation from monomers showed that calcium ions were more strongly bound to the α1(I) than the α2(I) chain. However, enhanced sampling methods implied the α2(I) chain has a higher trimer affinity than a third α1(I) chain in the presence of structural calcium. To quantify assembly thermodynamics, we computed relative binding free energies by alchemical thermodynamic integration, demonstrating that α2(I)-specific residues at the interchain interface conferred a measurable thermodynamic advantage to trimer formation in the presence of calcium. Hence, although heterotrimerization is normally favored, in reduced calcium conditions the homotrimer can form by sequestering available calcium to the α1(I) chains. This study provides a molecular explanation for a calcium-based mechanism driving heterotrimerization versus homotrimerization of type I collagen.