*Collagen: From Waste to Gold DOI: http://dx.doi.org/10.5772/intechopen.94266*

**Figure 2.**

*Process flow of transformation of hides into leather [12].*


#### **Table 1.**

*Chemicals used at each stage of hide to leather conversion and wastes generated [13].*

approximately 1000 amino acid residues. The accurate folding of these chains requires a glycine residue to be present in every third position of the polypeptide chain [1]. One-third of the amino acids in collagen in glycine and it always occupies the first position of the triplet. This is due to glycine being a small and an uncharged amino acid near the axis of the collagen triple helix. Glycine is a very crucial part of collagen molecule inherent characteristic as substitution of a single glycine for another amino acid disrupts the triple helix and results in skeletal deformities such as ontogenesis imperfect.

Imine acids make up approximately 25% of the residues in the collagen triplehelix. Imine acids – proline and hydroxyproline are typically found around the outside of the trip helix and the pentagon structure of these two amino acids includes the amine nitrogen and the α-carbon of the backbone chain. These limit the possible rotation in the amino acid (**Figure 1**) and hence forcing each collagen chain to form a left-handed helix. The high content of these imine acids makes the α-helix and β-sheet arrangements (generally found in proteins) unstable. Collagen triplehelix is held together by hydrogen bonding between chains. The NH group in

as fibres or sheets. A tropocollagen unit is about 285 kDa, 3000 Å in length and 15 Å in diameter. The triple helix is composed of repeating units of (Gly-X-Y)N amino acids, where X and Y are any amino acids, however, often X is proline and Y is hydroxyproline. The individual polypeptide chains of collagen each contain

*Process flow of waste valorisation from tanneries to collagen extraction and possible collagen-based applications.*

**Figure 1.**

*Biotechnological Applications of Biomass*

**206**

multiple types of hydrogen bonding patterns found in the triple-helix. These include, i) direct hydrogen bonding among the peptides (i.e. the NH group in glycine in each polypeptide chain forms H-bonds with adjacent peptide CO groups of the other chains), ii) water-mediated hydrogen bonding linking carbonyl groups, and iii) water-mediated hydrogen bonding, which links hydroxyproline OH groups and carbonyl groups. Collagen self-organisation forms bundles or a meshwork that determines the tensile strength and the elasticity and geometry of

The various collagen types are distinguished by the ability of their helical and non-helical regions to associate into fibrils and to form sheets or to cross-link different collagen types. For example, a two-dimensional network of type IV collagen is unique to the basal lamina. Most collagen is fibrillar and is composed of type I

Tropocollagen is produced by fibroblasts found in connective tissue in mammals and birds. The collagens α-chains are translated on the rough endoplasmic reticulum (RER). Inside the ER hydroxylation of the specific proline and lysine residues occurs, however lack of vitamin C will hinder this step. Inside the Golgi apparatus glycosylation of pro-α-chain lysine residues and formation of procollagen occurs. Procollagen molecules are exocytosed into extracellular space. The rest of the synthesis steps occur outside the fibroblasts. Procollagen peptidases cleave terminal regions of procollagen, transforming procollagen into insoluble tropocollagen. Many staggered tropocollagen molecules are reinforced by covalent lysinehydroxylysine cross-linkage (by Lysyl oxidase) to make collagen fibrils. Lysyl

*structure of collagen, with b) procollagen (loose ends), triple-helix wound together and c) collagen subunit tropocollagen (loose terminal removed) for final self-assemble of the collagen fibril and fibre (d-f) [21] (used*

the tissue.

**Figure 4.**

**209**

*with permission).*

molecules (**Figure 4**) [2].

*Collagen: From Waste to Gold*

*DOI: http://dx.doi.org/10.5772/intechopen.94266*

**3.2 Collagen synthesis**

oxidase requires copper (Cu++) for its activity [22].

#### **Figure 3.**

*Collagen structure being broken down to fibre, fibril, triple helix and an alpha chain respectively [20] (used with permission).*

glycine in polypeptide chains forms H-bonds with adjacent peptide CO groups of the other chains.

After the formation of the collagen polypeptide chain, proline in the third position of the triplet in the amino acid sequence is hydroxylated by the enzyme propyl hydroxylase. The hydroxyl groups of the hydroxyproline and water molecules form hydrogen bonds that stabilise the triple-helix. Inhibition of hydroxylation causes diseases such as scurvy (caused by a lack of vitamin C in the diet) which is the inability of the triple-helix to form at body temperature (37°C) [18]. A decrease in imine acids (proline and hydroxyproline) content lowers the thermal stability of collagen as collagen loses its helical structure and shrinkage or denaturation occurs [18]. Avian and mammalian collagen have very similar amounts of hydroxyproline at 13.5% of the total amino acids. In comparison, aquatic animals have a lower level of hydroxyproline at approximately 10.3% [19].

The alpha-triple helix of collagen is shaped into a right-handed helix. The alpha chains each are shaped into a left-handed symmetry (the opposite direction), and then three of these alpha coiled strands get together to form a right-handed triple helix so when under strain, the chains twist into each other, giving strength and preventing unravelling. Each alpha helix is approximately 1.4 nanometres in diameter and 300 nanometres in length (approx. 1000 amino acids). The collagen molecule can be composed of either three identical alpha chains (homotrimers), or two or three different alpha chains (heterotrimers), however, the chain configuration depends on the collagen type being synthesised [2]. The hierarchical structure of collagen is zoomed-in starting from the alpha chains coiling together to form the triple helix is shown in **Figure 3**.

Cross-links that are covalent bonds occur between the ends tropocollagen before the formation of the collagen fibre. The triple helix and the cross-linking give rise to a collagen material that is very rigid, inextensible and stable. Since collagen on the primary level is composed of repeating units of Gly-X-Y amino acids, it is therefore rich in carboxylic acid groups, hydroxyl groups, amide and amine groups. The triple helix structure is stabilised by inter-chain hydrogen bonding and triple helix (tropocollagen) molecules parallel to each other are covalently cross-linked with each other through their aldehyde and amino groups, forming collagen fibrils. There are

#### *Collagen: From Waste to Gold DOI: http://dx.doi.org/10.5772/intechopen.94266*

multiple types of hydrogen bonding patterns found in the triple-helix. These include, i) direct hydrogen bonding among the peptides (i.e. the NH group in glycine in each polypeptide chain forms H-bonds with adjacent peptide CO groups of the other chains), ii) water-mediated hydrogen bonding linking carbonyl groups, and iii) water-mediated hydrogen bonding, which links hydroxyproline OH groups and carbonyl groups. Collagen self-organisation forms bundles or a meshwork that determines the tensile strength and the elasticity and geometry of the tissue.

The various collagen types are distinguished by the ability of their helical and non-helical regions to associate into fibrils and to form sheets or to cross-link different collagen types. For example, a two-dimensional network of type IV collagen is unique to the basal lamina. Most collagen is fibrillar and is composed of type I molecules (**Figure 4**) [2].

#### **3.2 Collagen synthesis**

glycine in polypeptide chains forms H-bonds with adjacent peptide CO groups of

*Collagen structure being broken down to fibre, fibril, triple helix and an alpha chain respectively [20] (used*

After the formation of the collagen polypeptide chain, proline in the third position of the triplet in the amino acid sequence is hydroxylated by the enzyme propyl hydroxylase. The hydroxyl groups of the hydroxyproline and water molecules form hydrogen bonds that stabilise the triple-helix. Inhibition of hydroxylation causes diseases such as scurvy (caused by a lack of vitamin C in the diet) which is the inability of the triple-helix to form at body temperature (37°C) [18]. A decrease in imine acids (proline and hydroxyproline) content lowers the thermal stability of collagen as collagen loses its helical structure and shrinkage or denaturation occurs [18]. Avian and mammalian collagen have very similar amounts of hydroxyproline at 13.5% of the total amino acids. In comparison, aquatic animals

The alpha-triple helix of collagen is shaped into a right-handed helix. The alpha chains each are shaped into a left-handed symmetry (the opposite direction), and then three of these alpha coiled strands get together to form a right-handed triple helix so when under strain, the chains twist into each other, giving strength and preventing unravelling. Each alpha helix is approximately 1.4 nanometres in diameter and 300 nanometres in length (approx. 1000 amino acids). The collagen molecule can be composed of either three identical alpha chains (homotrimers), or two or three different alpha chains (heterotrimers), however, the chain configuration depends on the collagen type being synthesised [2]. The hierarchical structure of collagen is zoomed-in starting from the alpha chains coiling together to form the

Cross-links that are covalent bonds occur between the ends tropocollagen before the formation of the collagen fibre. The triple helix and the cross-linking give rise to a collagen material that is very rigid, inextensible and stable. Since collagen on the primary level is composed of repeating units of Gly-X-Y amino acids, it is therefore rich in carboxylic acid groups, hydroxyl groups, amide and amine groups. The triple helix structure is stabilised by inter-chain hydrogen bonding and triple helix (tropocollagen) molecules parallel to each other are covalently cross-linked with each other through their aldehyde and amino groups, forming collagen fibrils. There are

have a lower level of hydroxyproline at approximately 10.3% [19].

the other chains.

*Biotechnological Applications of Biomass*

*with permission).*

**Figure 3.**

triple helix is shown in **Figure 3**.

**208**

Tropocollagen is produced by fibroblasts found in connective tissue in mammals and birds. The collagens α-chains are translated on the rough endoplasmic reticulum (RER). Inside the ER hydroxylation of the specific proline and lysine residues occurs, however lack of vitamin C will hinder this step. Inside the Golgi apparatus glycosylation of pro-α-chain lysine residues and formation of procollagen occurs. Procollagen molecules are exocytosed into extracellular space. The rest of the synthesis steps occur outside the fibroblasts. Procollagen peptidases cleave terminal regions of procollagen, transforming procollagen into insoluble tropocollagen. Many staggered tropocollagen molecules are reinforced by covalent lysinehydroxylysine cross-linkage (by Lysyl oxidase) to make collagen fibrils. Lysyl oxidase requires copper (Cu++) for its activity [22].

#### **Figure 4.**

*structure of collagen, with b) procollagen (loose ends), triple-helix wound together and c) collagen subunit tropocollagen (loose terminal removed) for final self-assemble of the collagen fibril and fibre (d-f) [21] (used with permission).*
