A cysteine-based molecular code informs collagen C-propeptide assembly

Abstract

Fundamental questions regarding collagen biosynthesis, especially with respect to the molecular origins of homotrimeric versus heterotrimeric assembly, remain unanswered. Here, we demonstrate that the presence or absence of a single cysteine in type-I collagen’s C-propeptide domain is a key factor governing the ability of a given collagen polypeptide to stably homotrimerize. We also identify a critical role for Ca2+ in non-covalent collagen C-propeptide trimerization, thereby priming the protein for disulfide-mediated covalent immortalization. The resulting cysteine-based code for stable assembly provides a molecular model that can be used to predict, a priori, the identity of not just collagen homotrimers, but also naturally occurring 2:1 and 1:1:1 heterotrimers. Moreover, the code applies across all of the sequence-diverse fibrillar collagens. These results provide new insight into how evolution leverages disulfide networks to fine-tune protein assembly, and will inform the ongoing development of designer proteins that assemble into specific oligomeric forms.

Targeting defective proteostasis in the collagenopathies

“…The C-propeptide is a cysteine-rich domain whose folded state is stabilized by multiple intramolecular disulfide bonds. After the C-propeptide folds, individual C-propeptide domains recognize each other and assemble, a process that, at least for the fibrillar collagens, is mediated by Ca 2+ -and intermolecular disulfide bonds [62]. The resulting assembled C-propeptide trimer controls the triple helix register and initiates zipperlike folding of the proline-and glycine-rich triple-helical domain, a process that itself requires isomerization of hundreds of proline peptide bonds to the trans configuration.…”

Type III collagen (COL3A1): Gene and protein structure, tissue distribution, and associated diseases

“…The C-propeptides are important in the biosynthesis of fibrillar collagens by directing the chain selection, stabilizing the associated α chains by the formation of interchain disulphide bonds, and facilitating the formation of the triple helix (Boudko and Engel, 2004;Bourhis et al, 2012;DiChiara et al, 2018). In type III procollagen, the C-propeptide has eight cysteine residues and an N-glycosylation site.…”

Elucidation of proteostasis defects caused by osteogenesis imperfecta mutations in the collagen-α2(I) C-propeptide domain

“…Folded collagen-I C-Pro domains recognize appropriate partner C-Pro domains to nucleate the formation of 2:1 Cola1(I):Cola2(I) triple helices. This recognition process is mediated by a cysteine code that relies on a combination of Ca 21 -binding and intermolecular disulfide bond formation to stabilize the required 2:1 heterotrimers (10). Folding of the nearly 1000-amino-acid-long triplehelical domain then proceeds in the C !…”