Biomineralization and Nature’s Cryptic Mysteries: Collagens From Skin, Tendon, Dentin and Bone

Objectives: Objectives: Collagen type I is a major structural component of both mineralizing and non-mineralizing tissues such as bone, dentin, skin, and tendon. To clarify the ongoing debates regarding biomineralization mechanism, we have analyzed collagen type I from various tissue origin by state-of-the-art mass spectrometry (MS).
Methods: Methods: Collagen type I from mineralizing tissues such as bovine bone and from dentin of un-erupted molars were isolated in the laboratory and collagen type I from non-mineralizing tissues such as bovine tendon and skin were obtained from commercial sources. The complete topographical distribution of hydroxylation and phosphorylation states was identified and compared between collagen type I components derived from different tissue origin. The stable-isotope labelling chemistries were used for relative quantitative MS analysis which provided intricate details of the major similarities and distinctions between collagens derived from mineralizing versus non-mineralizing tissues.
Results: Results: Our qualitative studies indicated that collagen type I derived from mineralizing tissues such as bone and dentin contained 156 hydroxyproline sites from the total 175 proline residues in α-1 subunit, and 83 hydroxyproline sites from the total 180 proline residues in α-2 subunit. Importantly, there were 38 phosphorylation sites/α-1 subunit and 29 sites/ α- 2 subunit, with total of 105 phosphorylation sites per triple helix collagen molecule. The relative quantitative analysis indicated that collagen type I from mineralizing tissues were far superiorly phosphorylated in comparison to the non-mineralized tissue collagens. The alignment of the collagen molecules with specific phosphorylation regions highlighted very remarkable orientation and distribution of the phosphorylation sites. Each hole zone region would have up to 60 phosphorylation sites derived from two N-terminal and two C-terminal ends of four triple helix molecules.
Conclusions: Conclusions: The series of in-depth qualitative and relative quantitative MS analysis of collagen type I from mineralizing and non-mineralizing tissue origin revealed major structural differences with respect to phosphorylation states deemed to be critical in the normal mechanism of biomineralization and pathological conditions.