Structural and biochemical investigations of collagen-I trimerization

Abstract

The fibrillar collagens display a unique assembly process, with assembly initiating at the C-terminal propeptide (C-Pro) domain, a small globular domain that encodes vast amounts of information for chain selection, stoichiometry, and molecular recognition. The C-Pro domain is responsible for ensuring that strands that assemble together are of the same type of collagen, and, in the case of certain collagens, that the stoichiometry between strands is correct. In addition to being involved in assembly, the C-Pro domain has also been shown to play a significant role in collagen proteostasis. Quality control factors in the cell are able to specifically recognize misfolded C-Pro domains in the context of disease-causing mutations, demonstrating a vital role for C-Pro folding in this process. In chapter 2, we report progress towards the answer to a long-elusive question: why do certain collagens favor heterotrimeric assemblies even when homotrimeric assemblies are possible? We show progress towards a high-resolution structure of the collagen-I C-Pro 2:1 heterotrimer, and investigate the role of Ca2+ coordination in dictating assembly behavior and chain association. In chapter 3, we characterize the assembly and proteostasis defects associated with a set of C-Pro mutations that have been observed in patients with osteogenesis imperfecta. Our data reveal that, while some variants are effectively recognized by the cell’s quality control mechanisms and retained intracellularly, others are not and secrete at similar levels to the wild-type. We also demonstrate that even severely assembly-deficient C-Pro domains can escape quality control, likely resulting in severe or lethal disease. Collectively, the work presented here significantly advances our understanding of how collagen-I assembly occurs in healthy biological systems, and how it can be disrupted in disease, setting the stage for a variety of future investigations into this challenging system.