Structure and assembly of the heterotrimeric and homotrimeric C-propeptides of type I collagen: Significance of the α2(I) chain

Assembly of the type I procollagen molecule begins with interactions among the C-pro α1(I) and C-pro α2(I) domains. The C-propeptide domains themselves have subdomains of distinct structures. The important questions are where chain association begins and the basis of the chain selectivity which leads to the preferential formation of the [C-pro α1(I)]₂[C-pro α2(I)] heterotrimer. These questions are addressed by energy minimization modeling of the individual C-propeptide structures, study of their docking interactions, and comparison of the heterotrimeric and homotrimeric C-pro structures and stability. The comparisons show the remarkable impact of the C-pro α2 chain on the structure of the assembled trimeric C-propeptide. In the modeling, the three chains were anchored and registered by a short C-terminal collagen triple-helical segment followed by the C-telopeptides in their docked conformation, and then the remaining C-propeptide chains were allowed to interact and dock. Surprisingly, propeptide trimerization did not proceed through the previously proposed N-terminal "oligomerization domain" of the C-propeptide [McAlinden et al. (2003) J. Biol. Chem. 278, 42200] but rather in the most C-terminal domains of type I procollagen chains. Molecular dynamics showed heterotrimer assembly to begin with dimer formation between globular G2α2 and the G2α1₂ domains followed by trimerization at the G1 domains. Assembly initiation in the putative oligomerization coiled-coil domain is not possible because of the Pro residues at positions 3, 7, and 11 at the N-terminus of the α2 C-propeptide chain. To confirm the computations and proposed assembly pathway, the G2α1 and G2α2 domains were prepared recombinantly as the maltose binding protein constructs, and their interactions were studied by dynamic light scattering and gel filtration chromatography. Under the conditions examined MBP remained as monomer, MBP-G2α1 and MBP-G2α2 alone formed dimers, but a 2:1 mixture of MBP-G2α1 and MBP-G2α2 favored trimer formation. Thus, the C-terminal globular domains (G2) of the type I collagen C-propeptides play a crucial role in the initiation of intermolecular assembly and heterotrimer selectivity.