Collagen Marker: Hydroxyproline
by Eben van Tonder
20 July 2020
Many people drink collagen to maintain a youthful appearance of the skin and to provide support for tendons which may show signs of old age. The important part of the collagen protein they are relying on is hydroxyproline. Irrespective of the animal species, collagen fibres have an amino acid composition in which glycine makes up around one-third of the total residues and the amino acids proline and hydroxyproline a further 15-30%. “Hydroxyproline is of very limited occurrence in proteins, the only other mammalian protein in which it occurs being elastin (2 %). Collagen is also the only protein reported to contain more than about 0.1 % hydroxy-lysine.” (Courtis and Ward, 1977) The presence of hydroxyproline is the marker used to determine the approximate inclusion of collagen into meat products. On the other hand, too much hydroxyproline levels in one’s urine and/or serum are normally associated with degradation of connective tissue. Hydroxyproline can never be reincorporated into newly synthesized collagen, but both collagen synthesis and tissue breakdown contribute to urinary hydroxyproline. (Cremers, et al. 2008)
Looking at this matter is an interesting way of becoming more familiar with this fascinating molecule and how our body uses it in digestion.
Enzymes Responsible for its Formation
“4-Hydroxyproline, 3-Hydroxyproline, hydroxylysine are found in animals, almost exclusively in collagen. 4-Hydroxyproline is also found in plants. All three amino acids are formed as cotranslational and posttranslational modifications by the hydroxylation of peptide-bound proline or lysine residues by three separate enzymes, prolyl 4-hydroxylase, prolyl 3-hydroxylase and lysyl hydroxylase. Prolyl 4-hydroxylase is widely distributed in nature, as it has now been identified in a number of vertebrate, invertebrate and plant sources. The other two enzymes are also found in a variety of animal sources, but they appear to be absent from plants.” (Harding, et al. 1992)
“Hydroxyproline in protein, predominantly in collagen, is formed from proline after peptide linkage. Preformed hydroxyproline, whether derived from dietary protein or released from collagen turnover, is not incorporated into protein. Since the hydroxyproline produced from OH-P5C is not used for protein formation, we considered the possibility that hydroxyproline oxidase participates in redox generation. The role of the proline cycle in redox generation is well documented; therefore, the question arose whether the hydroxyproline cycle may play a similar role.” (Harding, et al., 1992)
“Prolyl 4-hydroxylase plays a central role in collagen synthesis as the 4-hydroxyproline residues formed in the reaction are an absolute requirement for the folding of the newly synthesized procollagen polypeptide chain into triple helix molecules. In plant proteins, the 4-hydroxyproline residues are frequently O-glycosylated and may form intermolecular cross-links which contribute to cell-wall assembly and stability. The hydroxylysine residues of animal proteins have two important functions: their hydroxyl groups serve as sites of attachment for carbohydrate units, and they are essential for the stability of certain collagen cross-links. The role of 3-hydroxyproline residues is unknown at present.” (Harding, et al. 1992)
In terms of its occurrence in collagen, “4-hydroxyproline content shows only small, though, distinct differences between collagens in most cases whereas 3-hydroxyproline and hydroxylysine varies markedly. In the case of 3-hydroxyproline, the value range from 0 to over 10 resides per 1000 amino acids, and in the case of hydroxylysine from about 5 to 70. Additional variations are found in the content of the three amino acids within the same collagen type in different tissues in many physiological and pathological states. The 4-hydroxyproline content of a given collagen type, again, only varies only within narrow limits, apparently because of the function of 4-hydroxyproline, whereas large variations are found in 3-hydroxyproline and hydroxylysine.” (Harding, et al. 1992)
Cooper SK, Pandhare J, Donald SP, Phang JM. 2008. A novel function for hydroxyproline oxidase in apoptosis through generation of reactive oxygen species. J Biol Chem. 2008; 283(16):10485-10492. doi:10.1074/jbc.M702181200
Cremers, S., Garnero, P., Seibel, M. J.. 2008. Chapter 87 – Biochemical Markers of Bone Metabolism. Principles of Bone Biology (Third Edition), Volume II, 2008, Pages 1857-1881
Harding, J. J., James, M. and Crabbe, C. 1992. Post-translational Modifications of Proteins. CRC Press.