scholarly journals Type-III procollagen assembly in semi-intact cells: chain association, nucleation and triple-helix folding do not require formation of inter-chain disulphide bonds but triple-helix nucleation does require hydroxylation

1996 ◽  
Vol 317 (1) ◽  
pp. 195-202 ◽  
Author(s):  
Neil J. BULLEID ◽  
Richard WILSON ◽  
Janice F. LEES
Keyword(s):  
Triple Helix ◽  
Intact Cells ◽  
Permeabilized Cells ◽  
Type Iii ◽  
Helical Domain ◽  
Helix Formation ◽  
Disulphide Bonds ◽  
Type Iii Procollagen ◽  
Triple Helix Formation ◽  
Triple Helical Domain

Procollagen assembly is initiated within the endoplasmic reticulum by three α-chains associating via their C-propeptides (C-terminal propeptides). To study the requirements for the association of procollagen monomers at synthesis we have reconstituted the initial stages in the folding, assembly and modification of procollagen using semi-permeabilized cells. By translating a type-III procollagen ‘mini-gene’ which lacks part of the triple-helical domain, we demonstrate that these cells efficiently carry out the assembly of hydroxylated, triple-helical, procollagen trimers and allow the identification of specific disulphide-bonded intermediates in the folding pathway. Mutant chains, which lack the ability to form inter-chain disulphide bonds within the C-propeptide, were still able to assemble within this system. Furthermore, characterization of the trimeric molecules formed suggested that inter-chain disulphide bonds had formed within the C-telopeptide (C-terminal telopeptide). However, when hydroxylation of prolyl and lysyl residues was inhibited no inter-chain disulphide bonds were formed in the C-telopeptide, indicating that hydroxylation is required for the initial nucleation of the triple-helical domain. Mutant chains which lacked the ability to form inter-chain disulphide bonds within the C-propeptide or the C-telopeptide could still assemble to form trimeric triple-helical molecules linked by inter-chain disulphide bonds within the N-propeptide (N-terminal propeptide). These results indicate that inter-chain disulphide bond formation within the C-propeptide or the C-telopeptide is not required for chain association and triple-helix formation.

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

2020 ◽  
Vol 295 (29) ◽  
pp. 9959-9973 ◽  
Author(s):  
Ngoc-Duc Doan ◽  
Azade S. Hosseini ◽  
Agata A. Bikovtseva ◽  
Michelle S. Huang ◽  
Andrew S. DiChiara ◽  
...  
Keyword(s):  
Quality Control ◽  
Endoplasmic Reticulum ◽  
Osteogenesis Imperfecta ◽  
Triple Helix ◽  
Helix Formation ◽  
Triple Helix Formation ◽  
Triple Helices ◽  
Triple Helical Domain ◽  
Key Aspects ◽  
Intracellular Collagen

Intracellular collagen assembly begins with the oxidative folding of ∼30-kDa C-terminal propeptide (C-Pro) domains. Folded C-Pro domains then template the formation of triple helices between appropriate partner strands. Numerous C-Pro missense variants that disrupt or delay triple-helix formation are known to cause disease, but our understanding of the specific proteostasis defects introduced by these variants remains immature. Moreover, it is unclear whether or not recognition and quality control of misfolded C-Pro domains is mediated by recognizing stalled assembly of triple-helical domains or by direct engagement of the C-Pro itself. Here, we integrate biochemical and cellular approaches to illuminate the proteostasis defects associated with osteogenesis imperfecta-causing mutations within the collagen-α2(I) C-Pro domain. We first show that “C-Pro-only” constructs recapitulate key aspects of the behavior of full-length Colα2(I) constructs. Of the variants studied, perhaps the most severe assembly defects are associated with C1163R C-Proα2(I), which is incapable of forming stable trimers and is retained within cells. We find that the presence or absence of an unassembled triple-helical domain is not the key feature driving cellular retention versus secretion. Rather, the proteostasis network directly engages the misfolded C-Pro domain itself to prevent secretion and initiate clearance. Using MS-based proteomics, we elucidate how the endoplasmic reticulum (ER) proteostasis network differentially engages misfolded C1163R C-Proα2(I) and targets it for ER-associated degradation. These results provide insights into collagen folding and quality control with the potential to inform the design of proteostasis network-targeted strategies for managing collagenopathies.

scholarly journals Effect of prevention of procollagen triple-helix formation on proline 3-hydroxylation in freshly isolated chick-embryo tendon cells

1981 ◽  
Vol 196 (1) ◽  
pp. 203-206 ◽  
Author(s):  
K Majamaa
Keyword(s):  
Carboxylic Acid ◽  
Chick Embryo ◽  
Triple Helix ◽  
Hydroxylase Activity ◽  
Intact Cells ◽  
Helix Formation ◽  
Tendon Cells ◽  
Triple Helix Formation

Inhibition of procollagen triple-helix formation by the addition of cis-hydroxyproline or azetidine-2-carboxylic acid increased the synthesis of 3-hydroxy[14C]proline 1.7-1.8-fold in pulse-chase experiments with freshly isolated chick-embryo tendon cells. The amount of 3-hydroxy[14C]proline, expressed as a percentage of the total 14C radioactivity in hydroxyproline, reached 8.4%. Control experiments indicated that the two analogues had no effect on the prolyl 3-hydroxylase activity of these cells. The data suggest that the time available before triple-helix formation in part regulates the extent of the 3-hydroxylation of proline in the biosynthesis of collagen in intact cells.

scholarly journals A physico-chemical study of triple helix formation by an oligodeoxythymidylate with N3'->P5' phosphoramidate linkages

1997 ◽  
Vol 25 (9) ◽  
pp. 1782-1787 ◽  
Author(s):  
B.-w. Zhou-Sun ◽  
J. Liquier ◽  
E. Taillandier ◽  
J.-s. Sun ◽  
T. Garestier ◽  
...  
Keyword(s):  
Triple Helix ◽  
Chemical Study ◽  
Helix Formation ◽  
Physico Chemical ◽  
Triple Helix Formation
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