Collagen synthesis-Dr.Bijay.pdf

Collagen Synthesis Dr. Bijay Barik Assistant Professor Dept. of Biochemistry, Cell Biology and Genetics Office: QC 22 [email protected] 1 MCB.10. Understand the structure, synthesi...

Collagen Synthesis Dr. Bijay Barik Assistant Professor Dept. of Biochemistry, Cell Biology and Genetics Office: QC 22 [email protected] 1 MCB.10. Understand the structure, synthesis, and distribution of collagen and its clinical significance. Given a clinical or experimental scenario or diagram, students should be able to: MCB.10.1. Differentiate collagen from other proteins with respect to the structural features and predominant amino acid composition. MCB.10.2. Distinguish the major collagen types and their distribution among tissues. MCB.10.3. Illustrate the steps and sites involved in the synthesis and processing of collagen. MCB.10.4. Describe the roles of vitamin C and copper in collagen synthesis and the clinical manifestations of their deficiency. Reference: Lippincott Illustrated Reviews, Biochemistry, 8th Edition (pg. 45-50) 2 3 4 MCB.10. Understand the structure, synthesis, and distribution of collagen and its clinical significance. Given a clinical or experimental scenario or diagram, students should be able to: MCB.10.1. Differentiate collagen from other proteins with respect to the structural features and predominant amino acid composition. MCB.10.2. Distinguish the major collagen types and their distribution among tissues. MCB.10.3. Illustrate the steps and sites involved in the synthesis and processing of collagen. MCB.10.4. Describe the roles of vitamin C and copper in collagen synthesis and the clinical manifestations of their deficiency. 5  Collagen and elastin are commonly occurring, well- characterized fibrous proteins of extracellular matrix (ECM) Both of these proteins serve a structural function in the body Collagen is the most abundant protein in the human body. A typical collagen molecule is a long rigid structure with three polypeptide chains (α-chains) These three α-chains wound around each other to form a rope-like triple helix structure. 6  The three α-chains of the triple helix are held together by interchain hydrogen bonds. Variations in the amino acid sequence of α chains result in different types of collagen with different properties 7  Salient features of collagen The 3 α - polypeptide chains form a triple helix Each chain is rich in amino acids glycine and proline Glycine the smallest amino acid (lack side chain), can fit into the restricted space where three chains of the helix come together. Each -chain has 1000 amino acids Every third position is occupied by Glycine Repeating sequence of ( -Gly- X-Y- )333 X- usually Proline Y- usually Hydroxyproline or hydroxylysine (post-translationally modified amino acids) 8  So, most frequently the sequence of the α-chain may be -(Glycine - proline – hydroxyproline/hydroxylysine)- Proline facilitates helical conformation of each α-chain because its ring structure causes “kinks” (sharp twist) in the peptide chain Hydroxyproline/hydroxylysine is not present in most other proteins Hydroxyproline maximizes the formation of interchain hydrogen bonds which stabilizes the triple helical structure. 33% glycine , 21% (proline + hydroxyproline) 9 MCB.10. Understand the structure, synthesis, and distribution of collagen and its clinical significance. Given a clinical or experimental scenario or diagram, students should be able to: MCB.10.1. Differentiate collagen from other proteins with respect to the structural features and predominant amino acid composition. MCB.10.2. Distinguish the major collagen types and their distribution among tissues. MCB.10.3. Illustrate the steps and sites involved in the synthesis and processing of collagen. MCB.10.4. Describe the roles of vitamin C and copper in collagen synthesis and the clinical manifestations of their deficiency. 10  Types of Collagen More than 25 types of collagen occur in human bodies - Type I, II, III, IV, and V etc. Most common collagen: Type I collagen (two α1 + one α2) Type II collagen – three α1 chains (α13) Collagen may be dispersed as gel (ECM), bundled into parallel fibers in tendons to provide great strength, or present as fibers arranged at an angle to each other (bones) to resist mechanical shear. Based on the location & functions collagens are organized into three different groups Type IV form a 3-dimensional mesh, and function as a semipermeable filtration barrier to macromolecules in the kidney and lungs 11 Types of Collagen (EM:Type IV polygonal network) 12 MCB.10. Understand the structure, synthesis, and distribution of collagen and its clinical significance. Given a clinical or experimental scenario or diagram, students should be able to: MCB.10.1. Differentiate collagen from other proteins with respect to the structural features and predominant amino acid composition. MCB.10.2. Distinguish the major collagen types and their distribution among tissues. MCB.10.3. Illustrate the steps and sites involved in the synthesis and processing of collagen. MCB.10.4. Describe the roles of vitamin C and copper in collagen synthesis and the clinical manifestations of their deficiency. 13 Biosynthesis of collagen: Synthesized in fibroblasts (connective tissue), osteoblasts (bone), and chondroblasts (cartilage) Collagen biosynthesis occurs as follows: 1. Formation of pro- chains 2. Hydroxylation 3. Glycosylation 4. Assembly and secretion 5. Extracellular cleavage of procollagen molecules 6. Formation of collagen fibrils 7. Cross-link formation 14 15 1. Formation of pro-α chains: ▪ In the nucleus, genes for prepro-α chains are transcribed (mRNA synthesis) ▪ mRNA is translated to prepro-α chain in Ribosome ▪ These Prepro-α chains contain a special amino acid sequence (signal sequence) at their N-terminal ends ▪ The signal sequence (hydrophobic amino acid rich sequence) facilitates the binding of ribosomes to the SRP (signal recognition particle) and this ribosome-peptide complex is recognized by SRP receptor present on rough endoplasmic reticulum (RER) and directs the passage of prepro-α chain into lumen of RER ▪ The signal sequence is then rapidly cleaved in the RER lumen (by signal peptidase) to yield a precursor of collagen called the Pro-α chain 16 2. Hydroxylation: The Pro-α chain undergoes hydroxylation in the RER Proline and lysine residues present in Y position (-Gly-X- Y-) of pro-α chain are hydroxylated by Prolyl hydroxylase & Lysyl hydroxylase, respectively. These hydroxylation reactions require ascorbic acid (vitamin C), Fe++ & molecular O2 3. Glycosylation: - Selected Hydroxylysine residues are then modified by glycosylation with glucose or glucosyl-galactose (in RER) 12 17  Vitamin C (Ascorbic acid) Vitamin C is a water-soluble vitamin. It is not stored in the body and excess amounts are excreted in the urine. Hence it must be supplied regularly in the diet to prevent deficiency. Functions of vitamin C include Important antioxidant Required for the synthesis of collagen (hydroxylation of proline and lysine) Enhances iron absorption Deficiency of Vitamin C: Scurvy Vitamin C is rich in fresh fruits and vegetables. Processed and canned food is low in vitamin C. Seen with “Tea and toast” diet (no fruits/vegetables) This disease is characterized by sore and spongy gums, loose teeth, fragile blood vessels, severe bruising, gingival bleeding, poor wound healing, poor bone mineralization, ecchymoses (bruise-like discoloration), and petechiae on limbs. Can also have swollen joints and fatigue. The observed symptoms in a patient suffering from scurvy may be due to a decrease in the hydroxylation process of collagen, resulting in the formation of defective collagen (less interchain H-bond and impaired triple helix) 18 Petechiae Subcutaneous leakage of Blood due to capillary fragility Ecchymoses 19 4. Assembly and secretion: - After hydroxylation and glycosylation, THREE Pro-α chains form Procollagen (precursor of collagen) - Procollagen has a central region of triple helix flanked by the non-helical N- and C- terminal extensions called Propeptides (Procollagen) Tropocollagen 20 - The formation of procollagen begins with formation of inter-chain disulfide bonds between the C-terminal extensions of pro-α chains - This brings the three α-chains into an alignment favorable for triple helix formation - The procollagen molecules move through the Golgi apparatus, where they are packaged in secretory vesicles - The vesicles fuse with the cell membrane, causing the release of procollagen molecules into the extracellular space (exocytosis)the 21 5. Extracellular cleavage of procollagen molecules: Once the procollagens are released into ECM, The triple helical procollagen molecules are cleaved by N- and C- procollagen peptidases → removal of disulfide-rich, N- and C-terminal propeptides → produce the Tropocollagen 6. Formation of collagen fibrils: Tropocollagen molecules spontaneously associate to form collagen fibrils. The fibrils form an ordered, parallel array, with adjacent collagen molecules arranged in a staggered pattern. 22 7. Cross-link formation Mature collagen fibers are formed by cross-link formation of individual fibrils The cross-link formation is facilitated by lysine & hydroxylysine of collagen First, some of the lysine & hydroxylysine residues are oxidatively deaminated by copper-containing enzyme Lysyl oxidase to form allysine and hydroxyallysine respectively (reactive aldehydes) Next, allysine & hydroxyallysine present in one collagen fiber condense with lysine & hydroxylysine of another collagen fiber to form covalent cross-links to form mature collagen fibers. Cross-linking increases with age 23  STAGGERED ARRANGEMENT OF COLLAGEN FIBRILS - Approximately three-quarters of each collagen fibril molecule overlaps the neighboring molecule. Collagen fibrils with cross-links forms collagen fiber with 24 Overview of Biosynthesis of Collagen 25 Degradation of collagen: Collagens are highly stable molecules Have relatively long half-lives of several years Breakdown of collagen fibers is dependent on the proteolytic action of Collagenases present in the matrix (matrix metalloproteinase) 26  Disorders of Collagen Biosynthesis Disease Defect Symptoms Scurvy Vitamin C deficiency, Loose teeth, bleeding gum, poor wound Hydroxylation reaction healing, ecchymoses, petechiae, poor bone defect development Menkes disease Deficient cross-link Depigmented (steely/kinky) hair, arterial formation, secondary to tortuosity, cerebral degeneration, lack of functional copper muscle growth, osteoporosis, anemia deficiency 27  Disorders of Collagen Biosynthesis Disease Defect Symptoms Ehlers-Danlos Deficiency of lysyl/prolyl Skin extensibility and fragility and joint syndrome hydroxylase or hypermobility procollagen peptidase or mutation in collagen gene Osteogenesis Collagen gene mutation Bone fragility, multiple fractures, hearing loss, imperfecta (Type I collagen defect) blue sclerae etc. Alport syndrome Mutation in type IV Defect in kidney basement membrane, collagen gene hematuria, proteinuria, hearing loss etc. Epidermolysis Mutation in genes for Skin becomes extremely fragile, easily bullosa collagen or laminin blistered and eroded (butterfly skin disease) 28 Ehlers-Danlos syndrome 29  Blue sclera Osteogenesis imperfecta 30 MCB.10. Understand the structure, synthesis, and distribution of collagen and its clinical significance. Given a clinical or experimental scenario or diagram, students should be able to: MCB.10.1. Differentiate collagen from other proteins with respect to the structural features and predominant amino acid composition. MCB.10.2. Distinguish the major collagen types and their distribution among tissues. MCB.10.3. Illustrate the steps and sites involved in the synthesis and processing of collagen. MCB.10.4. Describe the roles of vitamin C and copper in collagen synthesis and the clinical manifestations of their deficiency. 31 References: Lippincott’s Illustrated reviews : Biochemistry, 8th edition (pg. 45-50) 32 Please mail your questions to [email protected] 33