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. 2019 Apr 12;17(4):223.
doi: 10.3390/md17040223.

Isolation and Characterisation of Major and Minor Collagens from Hyaline Cartilage of Hoki (Macruronus novaezelandiae)

Mathew H Cumming  1 , Bronwyn Hall  2 , Kathleen Hofman  3
Affiliations

Isolation and Characterisation of Major and Minor Collagens from Hyaline Cartilage of Hoki (Macruronus novaezelandiae)

Mathew H Cumming et al. Mar Drugs. .

Abstract

The composition and properties of collagen in teleost (bony fish) cartilage have never been studied. In this study, we aimed to identify and characterise all collagen species in the nasal cartilage of hoki (Macruronus novaezelandiae). Four native collagen species were extracted using two techniques, and isolated with differential salt precipitation. We were able to assign the identity of three of these collagen species on the basis of solubility, SDS-PAGE and amino acid analyses. We found that hoki cartilage contains the major collagen, type II, and the minor collagens, type IX and type XI, which are homologous to those found in mammal and chicken cartilage. Using these extraction protocols, we also isolated a full-length type IX collagen from cartilage for the first time. In addition, we detected a 90 kDa, highly glycosylated collagen that has not been identified in any other species. For each isolate, structural and biochemical characterisations were performed using circular dichroism and Fourier transform infrared spectroscopy analyses, and the thermal denaturation properties were determined. Our results showed that the properties of hoki cartilage-derived collagens are similar to those of collagens in mammalian cartilage, indicating that teleost cartilage could provide biological ingredients for the development of biomaterials to treat cartilage-related illnesses.

Keywords: arthritis; cartilage; fish; marine; type II collagen; type IX collagen; type XI collagen.

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Conflict of interest statement

The authors declare no conflicts of interest.

Figures

Figure 1
Figure 1
(a) Nasal cartilage from hoki; (b) flow diagram for extraction of collagens from hoki cartilage.
Figure 2
Figure 2
SDS-PAGE analysis of differentially salt (NaCl)-precipitated collagens from hoki cartilage. Lanes 2–5, from pepsin-soluble fraction. Lanes 6–9, from alkaline-soluble fraction. Lanes 1 and 10, Broad range marker; lanes 2 and 6, proteins precipitated by 0.8 M NaCl; lane 3, proteins precipitated by 1.0 M NaCl; lane 7, proteins precipitated by 1.2 M NaCl; lanes 4 and 8, proteins precipitated by 2 M; lanes 5 and 9, same proteins as in lanes 4 and 8 after incubation with a reducing agent.
Figure 3
Figure 3
Circular dichroism wavelength spectra of (a) pepsin-soluble hoki samples: Type II collagen (━━), 90 kDa protein (━━), Type IX (━━); (b) alkaline-soluble samples: Type II collagen (━━), Type XI (━━), Type IX (━━).
Figure 4
Figure 4
Thermal melting curves of collagens extracted from hoki cartilage. Pepsin-soluble type II collagen (━━), type IX (━━), 90 kDa (━━). Alkaline-soluble type II collagen (┅), type XI (┅), type IX (┅).
Figure 5
Figure 5
Fourier transform infrared spectra of collagens isolated from hoki cartilage. Pepsin-soluble type II collagen (━━), type IX (━━), 90 kDa (━━). Alkaline-soluble type II collagen (━━), type XI (━━), type IX (━━).
Figure 6
Figure 6
Second derivative Fourier transform infrared spectra of collagens isolated from hoki cartilage between 1500 and 12,700 cm−1. Pepsin-soluble type II collagen (━━), type IX (━━), 90 kDa (━━). Alkaline-soluble type II collagen (━━), type XI (━━), type IX (━━).
Figure 7
Figure 7
Second derivative Fourier transform infrared spectra of collagens isolated from hoki cartilage between 825 and 1200 cm−1. Pepsin-soluble type II collagen (━━), type IX (━━), 90 kDa (━━). Alkaline-soluble type II collagen (━━), type XI (━━), type IX (━━).
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