Presentation on theme: "Specialized tissue Proteins:Collagen and Elastin"— Presentation transcript:
1Specialized tissue Proteins:Collagen and Elastin Dr.S.Chakravarty MBBS, MD
2Learning objectives:Describe the structure and formation of collagen and elastinList the various steps in Post translational modification of collagenMention the role of Vitamin C and copper in stabilizing the collagen structureList the types of collagen and its distribution in the bodyDescribe the defects of collagen and elastin and its associated clinical conditions
4Functions of extracellular matrix Regulation of proliferation, differentiation, migration and cell-cell recognitionPrevents or limits the movement of bacteria and cancer cellsDamage leads to various diseases like osteoarthritis, Glomerulonephritis etc.
5Property of fibrous proteins Alpha helical secondary structure.Low water solubilityA long narrow rod like structure.Role in determining cellular structure and function.
6Types of fibrous proteins: Collagen - most abundant protein in body; rigid, insoluble.Elastin - stretchy, rubber-like, lungs, walls of large blood vessels, ligamentsKeratin - tough fibers (hair, nails, outer epidermis)
7Collagen –many functions in many tissues !! Dispersed as a gel – Vitreous humorTight parallel fibres – TendonsStacked for minimal scattering – CorneaMechanical shearing – Bone.
8Types of collagen:So far, 28 types of collagen have been identified and described. The five most common types are:Fibril formingTissueFunctionType 1 (90%)Tendon, bone, ligaments and skinResistance to tensionType 2Hyaline and elastic cartilageResistance to pressureType 3Skin, muscle, blood vesselsStructural framework for expanding tissuesNetwork formingType 4Basement membraneFiltration and supportAnchoring fibrilsType 7EpitheliumAnchors basal cells to underlying stromaType I collagen is stronger than steel !!
10OutlineMolecular collagen (pre pro collagen and pro collagen) – solubleMicrofibrils – tropocollagen ( insoluble)FibrilsFibres
11Collagen triple helix Usmle! Left handed helix 3 such strands wound together
12About 25-30% of the total weight of body is collagen. Major fibrous element of tissues like bone , teeth , tendons , cartilage and blood vessels.Each polypeptide has about 1000 amino acid residues.1/3 of the a.a are Gly residues i.e every 3rd residue is glycine.The repetitive a.a sequences can be denoted by Gly-X-Y , where X and Y are commonly Proline and Hydroxyproline .
13The collagen is a rod like structure . The three polypeptide chains are held in a helical conformation by winding around each other.This results in formation of a superhelical cable with 3.3 amino acids per turn and each turn separated by 2.9 A.The strands are H-bonded to each other ( H-donated by NH grp and H-accepted by C=O )Further stabilization by H –bonds between OH- groups and the bridging water molecules.
16Quarter staggered Arrangement The trophocollagen molecules are arranged in in such a way that each row moves ¼ length over last row and the 5th row repeats the same position of the first row.Molecules in each row separated by 400 A and adjacent and adjacent rows by 680 A.The collagen fibres are further strengthened by covalent cross links b/w lysine and hydroxy-lysine
18Formation of pro alpha chains: Cytoplasm of fibroblastsFormation of pro alpha chains: with signal sequence at N-terminal ends.Rough-endoplasmic reticulum: signal sequence directs proteins to RER.Removal of signal sequence
19Events in the RER :Formation of pro-collagen Hydroxylation of proline and lysineRequires a dioxygenase with Fe .(Vit C keeps the iron reduced )Glycosylation – hydroxylysine with glucose.Spontaneous disulfide bond formation at C terminal peptidesformation of triple helix.3. Assembly in the Golgi and release of pro-collagen to extra cellular matrix.USMLE concept!
204. Extracellular matrix Stabilizing force H-bonding between Gly of one chain and Pro of another~1 H-bond per tripletExtra cellular cleavage of N and C-terminal propeptides– pro collagen peptidases.
21Tropo collagen molecules: Terminals (ends) of the triple-helix are differentC-telopeptidesN-telopeptidesTerminals are non-helicalHelps in triple helix formationN-TERMINAL INTRACHAIN DISULPHIDE BONDSC-TERMINAL- INTERCHAIN + INTRACHAIN DISULPHIDE BONDS(from Kadler, 1996)
22C-telopeptideElevated levels can be used in the confirmation of increased bone turnover.Elevated levels can identify persons with osteoporosis who have elevated bone turnover and who, as a result, are at increased risk for rapid disease progression.The patient's response to antiresorptive osteoporosis treatment can be monitored through this test.This test can be used to monitor and assess how effective antiresorptive therapy has been in patients treated for disorders such as osteopenia, osteoporosis, and Paget disease.This test can also serve as an adjunct means of monitoring patient response to other treatments for diseases with increased bone turnover, such as rickets & osteomalacia.
23Cross links formed by lysyl/prolyl oxidase copper co-enzymeOxidative deamination of lysines and hydroxylysines forms Allysine (aldehyde)This reacts with amino group of nearby lysine or hydroxylysine to form interchain cross-link.Very important for tensile strength of collagen.Cu2+/vitamin B6USMLE concept !
24Both extremes are bad !!Excessive cross links problem in OLD AGE Hardening of ligaments (STIFF)Prone to tearLess cross links Weak collagenMenke’s disease due to decreased Cu (discussed later)
29Elastin Helps in retaining the shape after stretching. Connective tissue protein.lungs, large blood vessels, elastic ligaments
30Elastic fibresOuter cover - Microfibrils containing fibrillin and microfibril associated glycoproteins (15%)Core of amorphous elastin –single polypeptide chain of 800 amino acids-85%Non-polar amino acids – gly, ala, val. Also rich in pro, lysine. ( no OH-proline or OH-lysine)
31Elastin 3D network of cross-linked polypeptides – (tropo elastin) cross links involve Lys and alLys –lysyl oxidase4 Lys can be cross-linked into desmosineDesmosines account for elastic propertiesDesmosine
32Elastin Structure and Function Elastin interconverts between a number of conformations, both disordered (upper two on left) and b-spiral (bottom left).After cross-linking, when elastin is stretched (or compressed) it is less stable and it returns to the disordered conformations.Beta spiral. It has hydrogen bonds, transient, but compared to alpha helix or beta sheet it is secondary structure. If this is relaxed, and this is an extended chain, this does not represent alpha helix, there is cross linking holding the chains together. If you stretch them, elongated form of individual elastin monomers, held together by cross link. If you release whatever stress created the elongated structure, it spontaneously goes back to relaxed structure, does this in order to maximize the entropy. Example of entropy driven reaction. You would have to put energy in, to get it to elongate, this is a measure of disorder, so this is a more ordered structure, because they are elongated and lined up. When you take away the stress that is holding it in this structure, goes back to optimize entropy. If you were to squash it, it can be made smaller, and it would take energy to do that, then when you release pressure, more ordered because in smaller volume, release and it would go back to relaxed state that is larger. This is reversible. Resilient, flexible.6
33Elastin Cross-linking Some lysine residues in elastin are deaminated and oxidized to the aldehyde level.They combine with each other and with other lysines to form lysinonorleucine and desmosine cross-linksCross linking derived from lysines. Two primary structures. Lyinonorleucine, two lycines contribute, one elastin a lysine in one elastin polymer and what was a lysine in the other. One of them has been oxidized and deaminated. This is derived from two lysines where one loses its amino group, form covalent linkage. Linkage between elastins is as strong as covalent peptide bonds. More complex structure, desmosin is made up of four contributing side chains, each was a lysine, only one has the nitrogens and the other is oxidized to aldehyde from, and forms cross linking reactions. Cartoon doesn’t show four connected but you canUSMLE concept !7
34Table 48–5. Major Differences Between Collagen and Elastin Collagen ElastinMany different genetic types One genetic type2. Triple helix No triple helix; random coilconformations permitting stretching3. (Gly-X-Y)n repeating structure No (Gly-X-Y)n repeating structure4. Presence of hydroxylysine No hydroxylysine5. Carbohydrate-containing No carbohydrate6. Intramolecular aldol cross-links Intramolecular desmosine cross-links7. Presence of extension peptides No extension peptides present duringbiosynthesis during biosynthesisTable 48–5. Major Differences Between Collagen and ElastinCollagenElastin1. Many different genetic typesOne genetic type2. Triple helixNo triple helix; random coil conformations permitting stretching3. (Gly-X-Y)n repeating structure No (Gly-X-Y)n repeating structure 4. Presence of hydroxylysineNo hydroxylysine5. Carbohydrate-containingNo carbohydrate6. Intramolecular aldol cross-linksIntramolecular desmosine cross-links7. Presence of extension peptides during biosynthesisNo extension peptides present during biosynthesis
35Degradation of elastin: Serine type elastase: neutrophils, macrophages, fibrblasts.Matrix metalloproteinases – mmp-12 and 7, gelatinases.
36Keratin Keratin is rich in cysteines. Its secondary structure is mostly a-helical.The helices form coiled coils (on right).The coiled coils pack into higher order elongated structures.Keratin properties depend strongly on the degree of disulfide cross-linking.With low levels of cross-linking, it is flexible (hair, skin).It can be made very hard with additional cross-linking (claws, horns).2 nmThis lecture from structure function point of view, understand why some function occurs as a result of the structure of the molecules. The three examples, elastin, keratin, and collagen, have macroscopic properties that are directly related to the actual molecular structure. In the case of elastin, that structure was this spiral, random, coil type of structure, stretch and it springs back. Keratin is rich in cystines, almost all alpha helical. It forms coiled coils. this is similar to the tail region of myosin, two alpha helices that coil around one another. Coiled coils pack into more elaborate structures. There is cross linking, disulfide, between the chains of a single keratin structure and also between neighboring keratin structures that exist in higher order. Keratin is the main constituent of hair and nails, as well as the nails, there is a wide variation in how hard those different materials are. Keratin is also in skin, skin and hair are flexible, soft, claws and horns are not. The way that hardness is modulated is degree of cross linking. More cross links, harder the material. It results from cells being packed with keratin, cell when it is still alive makes keratin until its filled and then it dies but the cell stays in place filled with keratin connected to other cells. This structure is also occurring in other cases, like intracellular intermediate filament.8
37Keratin Supramolecular Structure Two coiled coils bind together to form a protofibril (below).Protofibrils assemble into various microfibrils (on the right).¬Fig. 4-5Fig. 4-6®RawnThe keratins form a protofibril when two of them form a complex that has a helical. Cross section is the two strands of alpha helix that make up keratin, binding to one another to form protofibril, four individual alpha helices, cross section of protofibril, four pink and orange circles. Protofibrils further aggregate to microfibrils. Two major microfibril arrangements are the square and the hexagonal, round shape. They are hollow inside, cross linking is along the length of the protofibril, everything is cross linked, with some there is enough resilience to regain its original structure. If you have a long hair and you can take some of it and pull on it, if you don’t pull too hard it stretches then goes back to original shape. Stretching alpha helices and when you let go it goes back to alpha helical form, guided back to that structure by sulfhydril cross linking. More cross linking, can bend finger nails.9
38Keratin Cross-linking The structure of keratin is strengthened by disulfide cross-links from one helix to another.Cross linking is disulfide cross link. You can make and break these cross links in the case of hair in reversible fashion. You can do that by manipulating this reaction, one method is to use heat, get the hair in the shape you want. Idea that you get it in that shape, heat it up, break disulfide bonds, then they reform and it maintains the structure for a while. Only form some of them. Another way to do it is chemically, chemicals break the bonds then other chemicals to form them, called a perm. Too much heat or chemical and it falls out. These are the keratin cross links.10
42MCQ 1A culture of fibroblast cells is provided with equal all the 20 amino acids. After 10 days , the concentration of the amino acids is assessed .Which amino acid will have the lowest concentration?A. LysineB.methionineC.GlycineD.prolineE.Cysteine
43Elastin fibres in the alveolar walls of the lungs can be stretched easily during inspiration and recoil to their original shape once the force is released. This process facilitates expiration. The property described can be best explained by:Heavy posttranslational hydroxylationHigh content of polar amino acidsChain assembly to form a triple helixInterchain crosslinks involving lysineAbundant interchain disulfide bridges
443. A 14-year old male presents to your office complaining of easy bruising. Physical examination reveals soft and loose skin as well as multiple ecchymoses in the forearm and pretibial regions. Histologic evaluation with electron microscopy shows collagen fibrils that are abnormally thin and irregular. Which of the following stages of collagen synthesis is most likely impaired in this patient?RNA signal sequence recognitionAmino acid incorporation into polypeptide chainTriple helix formationLysine residue hydroxylationcleavage of propeptides