Why study tissue proteins?

To understand normal body functions . In development In inflammatory states Aging process.

To identify the cause of various genetic and metabolic disorders related to tissue proteins In the spread of cancer cells. Several diseases (eg: Osteogenesis imperfecta and a number of

types of the Ehlers-Danlos syndrome) are due to genetic disturbances of the synthesis of collagen.

Components of proteoglycans are affected in the group of genetic disorders known as the mucopolysaccharidoses.

To use and apply them in the medical, industrial, commercial fields. In food products, cosmetic surgery. The gelatin used in food and industry is derived from the partial

hydrolysis of collagen.

Present in all organs (50 % of body weight) •skin, •adipose tissue,•bones, •teeth, •fascia,•cartilages, •stroma of parenchymal inner organs, •walls of vessels.

Cells

Fibers

Extracellular matrix

THE STRUCTURE OF CONNECTIVE TISSUE

Cells•Fibroblasts•chondroblasts

Fibers •collagen•elastin

Extracellular matrixcarbohydrate-protein complexes - proteoglycans. Carbohydrate complexes of proteoglycans – heteropolysaccharides glycosaminoglycans (mucopolysaccharides).

•fibroblasts

chondroblasts

The Extracellular Matrix The space outside the cells of a tissue is filled with a composite material

called extracellular matrix(ECM). This ECM is also known as the connective tissue. It is composed of -

a) Gel with interstitial fluid.

b) 3 major classes of biomolecules: Structural proteins: collagen, elastin and Keratin (epidermal tissues) Specialized proteins: e.g. fibrillin, fibronectin, and laminin. Proteoglycans(Mucoproteins) : Conjugated proteins consisting of

Protein + Carbohydrate(5%-95%) Carbohydrate part is in the form of Glycosaminoglycans [GAGs].

Thus, the extracellular matrix (ECM) is a complex structural entity surrounding and supporting cells that are found within mammalian tissues.

The Extra cellular matrix

Most abundant insoluble fibrous protein in the connective tissue of mammals. Makes up about 25% to 35% of the whole-body protein content. Scleroprotein secreted from the cells called fibroblasts. In greek ‘kolla’ means ‘glue’.Collagen is also called as glue-producer. Distribution of collagen varies in different tissues.

Also found in mucous membranes, nerves, blood vessels, and organs.

Collagen fibers in muscle tendons

PROTEINS OF CONNECTIVE TISSUE

Collagen

Functions Of Collagen

It imparts strength, support, shape and elasicity to the tissues. It accounts for 6% of the weight

of strong, tendinous muscles It provides flexibility, support, and

movement to cartilage. It encases and protects delicate

organs like kidneys and spleen. It fills the sclera of the eye in

crystalline form. Teeth (dentin) are made by adding

mineral crystals to collagen. Collagen contributes to proper

alignment of cells for cell proliferation and differentiation.

When exposed in damaged blood vessels, it initiates thrombus formation

Types Of Collagen In humans, there are at least 19

distinct types of collagen made up of 30 distinct polypeptide chains (each encoded by a separate gene).

They are subdivided into a number of classes based primarily on the structures they form

However, 90% of the collagen in the body are of type I, II, III, and IV.

These types determine the physical properties of specific tissues and perform their specialized function.

•It serves to hold together the cells in the tissues.

•It is the major fibrous element of tissues like bone, teeth, tendons, cartilage and blood vessels. •When a solution of collagen is boiled, the viscosity of the solution decreases, which indicates that the native rod like structure is altered and a protein, with random coil structure results. It is then called gelatin.

Collagen

Structure of Collagen

•There are 6 types of collagen, out of which type I is the most abundant form; it contains 2 chains of alpha-1 and one chain of alpha-2. •Each polypeptide chain of collagen has about 1000 amino acid residues.

•The amino acid composition of collagen is quite unique.

Consists of three polypeptide chains having the left spiral shape

Three left coiled chains are again coiled together to form the right spiral bunch

Structure of Collagen

1 chain contains about 1000 amino acids33 % - glycine21 % – proline and oxiproline11 %– alanine35 % – all other amino acids

Oxiproline and oxilysine are specific only for connective tissue Oxiproline

oxilysine

Structure of collagen fibers

Collagen – complex protein, glycoprotein

Carbohydrates (monosaccharide galactose and disaccharide galactosylglucose) bind by glycosidic bonds to the residues of oxilysine of polipeptide chain

Reaction of aldoll condensation

Biosynthesis of Collagen Collagen synthesis occurs in the fibroblasts, osteoblasts in bone,

chondroblasts in cartilage and odontoblasts in teeth. First synthesized in precursor form of preprocollagen polypeptide chain

in the ribosomes during translation The leader sequence of amino acids[signal peptide] in the

preprocollagen directs it to enter the lumen of E.R In the lumen of E.R, the Signal peptide is cleaved to form procollagen. The proline and lysine amino acids in the procollagen chain undergo

hydroxylation and glycosylation known as post translational modifications.

Disulfide bonds are formed between three procollagen chains which twist around each other to form a triple helix molecule.This step is called registration.

This Procollagen molecule is secreted into the extracellular matrix from the golgi compartment of the E.R.

Here, the procollagen aminoproteinase and carboxy proteinase enzymes remove extra terminal amino acids from the procollagen molecule to form collagen .

The collagen molecules assemble into fibrils and inturn fibers being stabilized by the covalent cross-links.

Biosynthesis of collagen from Preprocollagen

Synthesis Of Collagen

Abnormalities associated with collagen

Collagen-related diseases arise from genetic defects nutritional deficiencies

They affect the biosynthesis, assembly, postranslational modification, secretion, or other processes involved in normal collagen production.

These include : Ehler Danlos syndrome Alport syndrome Epidermolysis bullosa Osteogenesis Imperfecta Chondrodysplasias [affects cartilage] Scurvy Osteolathyrism

Ehler Danlos Syndrome Ehlers-Danlos Syndrome is a group of inherited connective tissue disorders. CAUSE

abnormalities in the synthesis and metabolism of collagen Mutations in the collagen genes: COL1A1, COL1A2, COL3A1, COL5A1, COL5A2 a deficiency of enzyme lysyl hydroxylase. A deficiency of procollagenN-proteinase, causing formation of abnormal thin,

irregular collagen fibrils

EFFECT Mutations alter the structure, production, or

processing of collagen or proteins that interact with collagen

WeakenS connective tissue in the skin, bones, blood vessels, and organs causing-

Skin hyperextensibility Joint dislocations Tissue fragility Poor wound healing.

Ehler-Danlos Syndrome

•Hyperextensibility of skin

•Hypermobility of joints

Alport Syndrome Alport syndrome is a genetic disorder

characterized by glomerulonephritis, endstage kidney disease, and hearing loss. It also affects the eyes. The presence of blood in the urine

[hematuria] is almost always found in this condition.

CAUSE Mutations in COL4A3,COL4A4,COL4A5

collagen biosynthesis genes. These prevent the production or

assembly of the type IV collagen network in the basement membranes.

kidneys are scarred and unable to filter waste products resulting in hematuria and renal disease.

Alport syndrome affecting eyes

Epidermolysis Bullosa• Epidermolysis bullosa refers to a group of

inherited disorders that involve the formation of blisters following trivial trauma.

• CAUSE mutations in COL7A1, affecting the

structure of type VII collagen. Type VII collagen forms delicate fibrils that

anchor the basal lamina to collagen fibrils in the dermis.

These anchoring fibrils are reduced in this form of the disease, causing friction and blistering.

• EFFECT Blistering and painful sores like third degree

burns

Blister formation

Osteogenesis Imperfecta Osteogenesis imperfecta or Brittle Bone

Disease is a genetic bone disorder due to decrease dcollagen formation.

CAUSE Mutations in the COL1A1 andCOL1A2

genes coding for procollagen chains. Replacement of glycine by another

bulkier amino acid resulting in decreased collagen or improper procollagen structure forming abnormal fibers.

Mutations also cause ‘procollagen suicide ‘

All these cause brittleness. EFFECT

Thin,t ransclucent, blue scleras. Affected infants may be born with

multiple fractures and not survive. weak muscles, brittle teeth, a curved

spine and hearing loss.

Chondrodysplasias Chondrodysplasias are a mixed

group of hereditary disorders affecting cartilage.

One example is Stickler syndrome, manifested by degeneration of joint cartilage and of the vitreous body of the eye.

CAUSE Mutations in the COL2A1

gene, leading to abnormal forms of type II collagen.

EFFECT shortlimbed dwarfism skeletal deformities.

Osteolathyrism Osteolathyrism is a collagen cross-linking

deficiency caused by dietary over-reliance on the seeds of Lathyrus sativus (kesari dal) in some parts of India.

CAUSE Osteolathyrogenic compounds like Beta-

aminopropionitrile(BAPN) and Beta-oxalyl aminoalanine [BOAA] found in Kesari dhal inhibit enzyme lysyl oxidase required for the formation of cross links in the triple helices

EFFECT weakness and fragility of skin, bones,

and blood vessels Paralysis of the lower extremities

associated with neurolathyrism

Scurvy Scurvy is a disease due to deficiency of

vitamin C It is not a genetic disease. It is related to improper collagen

formation CAUSE

Vitamin C [ascorbic acid ]is required as a cofactor for hydroxylase enzymes during the hydroxylation of proline and lysine in the synthesis of collagen.

Deficiency causes impaired collagen synthesis due to deficiency of hydroxylases.

EFFECT Bleeding of gums Poor wound healing Subcutaneous hemorrhages

Uses Of Collagen Industrial Uses

Collagen is used as temporary thermoplastic glues in musical instruments like violin and guitar .

Recently used as a fertilizer Gelatin derived from the partial hydrolysis of collagen is

used in food products like desserts, jellies. It is also used in pharmaceutical, cosmetic, and

photography industries. Medical uses

Mild benefit to rheumatoid arthritis patients. Keeps the valvular leaflets of heart in shape. Helps in the deposition of calcium during aging. Used in cosmetic surgery, for burn patients for

reconstruction of bone and a wide variety of dental, orthopedic and surgical purposes.

Main ingredient of cosmetic makeup. Human collagen is used for immunosuppression during

transplantation.

Elastin

The main constituent of elastic fibers in ligaments, walls of large arteries, lungs.

Molecule contains about 800 amino acid residues

Has globular shape.

Is joined into the fibrous cords.

Contains a lot of glycine, alanine, proline, valine

There are no oxilysine and cysteine

Residues of lisine form the cross covalent bonds

The net structure is formed which can strech two and more times.

PROTEOGLYCANS

Proteoglycans – the main extracellular matrix of connective tissue. Consist of protein part + polisaccharide chains Molecular weight – tens millions.

Polisaccharides - glycosaminoglycans (acidic mucopolisaccharides) are built from the large amount of identical disaccharide units.

Disaccharide – aminosugar N-acetylglucosamin or N-acetylgalactosamin + uronic acid (glucuronic or iduronic) + sulfate (sometime).

•Is contained in synovial fluid (lubricant in joints), vitreous substance of eye •In rheumatic diseases and arthritis hyaluronic acid is depolimerized and the viscosity of synovial fluid is decreased•Forms the viscous solutions•Retains water

Hyaluronic acid

Heparin

•Is synthesized by tissue basofils•During degranulation is ejected into the extracellular matrix•Participates in the regulation of blood coagulation. •Increases the release of enzyme lipoprotein lipase into blood plasma

Keratins

•Keratine are fibrous proteins present in hair, skin an nail, horn, hoof, etc.

•They mainly have the alpha helical structure. •Each fibril has 3 polypeptide chains and each bundle has about 10-12 fibrils.

• The matrix has cysteine-rich polypeptide chains which are held together by disulfide bonds. •The more the number of disulfide bonds, the harder the keratin is.